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209716123 | Biological activity assays We designed and synthesized more than 30 derivatives of DCZ0358. However, the ve-ring quaternary ammonium component of some derivatives was unstable to decompose easily into its hydrolyzate 23b. With 12 stable compounds in hand, we conducted experiments to assay inhibitory activities against both hCE1 and hCE2 using a panel of uorescent probe substrates. 33-36 D-Luciferin methyl ester (DME) was used as a probe substrate, and nevadensin (a specic hCE1 inhibitor) was used as a positive inhibitor control for hCE1. Fluorescein diacetate (FD) was used as a specic probe substrate, and loperamide (LPA) was used as a positive inhibitor control of hCE2. The IC 50 values of all derivatives were evaluated and listed in Table 1. Table 1 showed that the inhibitory effects of these compounds against hCE2 were enhanced signicantly when the methylenedioxy group on A ring was changed into benzyloxy group. with electron-donating groups on the benzyloxy ring were similar to that of 23e (hCE2 IC 50 11.46 AE 1.76 mM), 23f (hCE2 IC 50 5.73 AE 0.79 mM) and 23h (hCE2 IC 50 3.32 AE 0.87 mM) with electronwithdrawing groups. In terms of the selectivity, it improved apparently according to the values of IC 50 (hCE2)/IC 50 (hCE1) shown in Table 1. For instance, the value of IC 50 (hCE2)/IC 50 (hCE1) of 23o was up to 83 while that of 23a was only 0.25. Thus, 23o have the best selectivity on hCES2 among all these newly synthesized compounds. Collectively, the structure-activity relationships of these compounds were summarized as follows, (1) the oxazolinium moiety is crucial for the inhibitory activity against hCEs; (2) the benzyloxy group on the A ring mainly contributed to the selectivity of hCE2 over hCE1 (Fig. 3). The inhibition kinetic of 23o against hCE2-mediated FD hydrolysis has been carefully investigated and the results showed that 23o functioned as a mixed inhibitor against hCE2mediated FD hydrolysis, with the K i value of 0.84 mM (Fig. 4B). Furthermore, in view of that hCE2 is an intracellular enzyme, the inhibition potential of 23o was also investigated. As shown in Fig. 5, 23o could strongly inhibit intracellular hCE2-mediated NCEN hydrolysis and reduce the uorescence intensity in the green channel (for the hydrolytic metabolite of NCEN) in living HepG2 cells via a dose-dependent manner. Meanwhile, the IC 50 value of 23o against intracellular hCE2 was also evaluated as 2.29 mM (Fig. S2B †). | 4 |
209716123 | Molecular docking In order to investigate the interaction mechanism of 23o with hCE2, molecular docking of 23o to the active site of hCE2 was performed. As shown in Fig. 6, there are hydrogen bond between the methoxyl of ring D with Arg-355 (3.16Å), and a Ttype p-p interaction between the ring D with the Arg-355, as well as, hydrogen bond between the oxygen atom of ring B with Phe-307 (3.17Å) in the entrance of the active cavity of hCE2. These interactions facilitate the entry of 23o into the active cavity of hCE2. However, the hydrolysate of 23o cannot enter the active cavity of hCE2, due to its small inlet. In addition, the methoxyl group at the benzyloxy end of 23o could tightly bind to the catalytic amino acid Ser-228 (1.6Å) via strong H-bonding, as well as, with Ala-150 (3.18Å), and there are strong hydrophobic interactions between the benzyloxy group of 23o with the key residues in the active cavity of hCE2. These interactions may account for the high selectivity of 23o on hCE2. The strong Hbond interaction between 23o and Ser-228 indicates that 23o may obstruct hCE2-mediated hydrolysis, possibly because Ser- 228 is an important residue involved in substrate recognition and catalysis of hCE2. These ndings agreed well with the experimental data where 23o exhibited much more potent inhibitory effect on hCE2 but a relatively weaker one on hCE1. | 5 |
209716123 | Chemical synthesis Materials. All starting materials were obtained from commercial suppliers and used without further purication. The 1 H and 13 C NMR spectra were taken on Bruker Avance-600 or 500 or 400, Varian MERCURY Plus-400 or 300 NMR spectrometer operating at 400 MHz or 300 MHz for 1 H NMR, 125 MHz or 100 MHz for 13 C NMR, using TMS as internal standard and CDCl 3 or methanol-d 4 or DMSO-d 6 as solvent. 13 C NMR spectra were recorded with complete proton decoupling. The ESI-MS or EI-MS was recorded on Finnigan LCQ/DECA or Thermo-DFS, respectively. The HRMS were obtained from Micromass Ultra Q-TOF (ESI) or Thermo-DFS (EI) spectrometer. Flash column chromatography was carried out using silica gel (200-400 mesh). Thin layer chromatography (TLC) was used silica gel F254 uorescent treated silica that were visualised under UV light (254 nm). Synthetic procedure. Compounds DCZ0358 and 23b have been reported in our previous work. 18,20 Synthesis of 3,9,10-trimethoxy-5-(4-methoxy-3-((4-methoxybenzyl)oxy)phenyl)-2,3dihydrooxazolo[2,3-a]isoquinolin-4-ium (23d). To a solution of 22d (56 mg, 0.1 mmol) in acetone (5 mL) was added hydrochloric acid (1 mL, 2.0 M in diethyl ether), and then the mixture was stirred for 2 h at room temperature. The solution was evaporated in vacuo to obtain the titled compound 23d as yellow solid (46 mg, 83%). 1 161.2, 159.7, 152.9, 149.9, 147.3, 146.2, 136.8, 134.5, 130.9, 130.1, 126.1, 124.9, 124.5, 120.9, 118.8, 116 3,9,10-Trimethoxy-5-(4-methoxy-3-((4-(methylsulfonyl)benzyl) oxy)phenyl)-2,3-dihydrooxazolo[2,3-a]isoquinolin-4-ium (23e). Compound 23e was prepared from compound 22e (58 mg, 0.1 mmol) as a yellow solid (48 mg, 86%). 1 MD, USA) and stored at À80 C until use. DMSO was purchased from sher. Phosphate buffer was prepared using Millipore water and then stored at 4 C until use. All tested compounds were solved by DMSO and stored at 4 C until use. LC grade acetonitrile and DMSO (Tedia, USA) were used throughout. hCE1 inhibition assay. DME was used as a probe substrate for evaluating the inhibitory effects of all DCZ0358 derivatives on hCE1, while nevadensin (a specic hCE1 inhibitor) was used as a positive control. 39 Briey, 100 mL incubation mixture containing 91 mL PBS (pH 6.8), 2 mL inhibitor at different concentrations and 5 mL HLM (1 mg mL À1 , nal concentration), were pre-incubated at 37 C for 10 min. Subsequently, 2 mL DME (3 mM nal concentration, close to the K m value of DME in HLM) was added to initiate the reaction. Aer incubating for 10 min at 37 C in a shaking bath, the reaction was stopped by the addition of LDR (100 mL). The microplate reader (SpectraMax® iD3, Molecular Devices, Austria) was used for luminescence measurements. The gain value of luminescence detection was set at 140 volts, and the integration time was set at 1 s. The chemical structure of DME and its hydrolytic metabolite (Dluciferin), as well as the detection conditions for D-luciferin are depicted in Table S1. † The negative control incubation (DMSO only) was carried out under the same conditions. The residual activity was calculated using the following formula, the residual activity (%) ¼ (the orescence intensity in the presence of inhibitor)/the orescence intensity in negative control  100%. The residual activities are show in Fig. S1. † hCE2 inhibition assay. The inhibitory effects of all DCZ0358 derivatives on hCE2 were investigated using uorescein diacetate (FD) as a specic probe substrate, 40 while LPA was used as a positive inhibitor of hCE2 in this study. 41 In brief, 200 mL incubation mixture containing 0.1 M PBS (PH ¼ 7.4), human liver microsomes (2 mg mL À1 , nal concentration) and each inhibitor. Aer 10 min pre-incubation at 37 C, the reaction was initiated by adding FD (5 mM, nal concentration, close to the K m value of FD in HLM). Aer incubating for 30 min at 37 C in a shaking bath, the reaction was stopped by the addition of acetonitrile (200 mL). The chemical structure of FD and its hydrolytic metabolite (uorescein), as well as the detection conditions for uorescein are depicted in Table S1. † The negative control incubation (DMSO only) was also carried out under the same conditions. 42 The residual activity was calculated using the formula mentioned above in hCE1 inhibition assay. The residual activities are shown in Fig. S1. † Cell culture and uorescence imaging analyses. In view of that hCE2 was an intracellular enzyme, the inhibition potential of 23o was investigated in living HepG2 cells. The HepG2 cells were cultured in Modied Eagle's Medium (MEM) with 5% CO 2 and 0.1% antibiotic-antimycoticmix antibiotic at 37 C, supplemented with 10% fetal bovine serum (FBS) and used NCEN as substrate probe to assay the 23o inhibition potential toward hCE2. NCEN, 43 another specic optical probe substrate for hCE2, the structure and hydrolytic site were shown in Fig. S2(A). † For uorescence imaging, HepG2 cells were seeded in 96well plates (8000 cells per well) with complete medium and then incubated for 24 hours. Aerwards, the cells were washed twice with FBS-free culture medium and then preincubated in the medium containing 23o (prepared in FBS-free at various concentrations) for 30 min with 5% CO 2 at 37 C. HepG2 cells were then co-incubated with NCEN (nal concentration, 10 mM) for another 50 min to assess the intracellular hCE2 function, respectively. The living cells were imaged and analyzed using an ImageXpress® Micro Confocal High-Content Imaging system (Molecular Devices, Austria). | 7 |
99398003 | A novel bioinspired molecule, 1,3-bis(vinylbenzyl)thymine (bisVBT), was isolated as a by-product during the synthesis of 1-(4vinylbenzyl)thymine (VBT) and analyzed with various techniques: NMR, IR, and Single-Crystal X-ray Diffraction. In addition to embodying all the desired characteristics of VBT (i.e., having the ability to undergo a 2π + 2π photodimerization reaction upon UV irradiation, a derivatization site, hydrogen bonding sites, and aromatic stacking ability) the bisVBT monomer has the added benefit of having two vinyl groups for cross-polymerization. Copolymerizing the bisVBT monomer with the charged monomer vinylbenzyl triethylammonium (VBA) chloride, different copolymers/terpolymers/cross-linked network were obtained, as it was shown by the absence of the vinyl resonance in the NMR spectra. Thermal Gravimetric Analysis (TGA) and Differential Scanning Calorimetry (DSC) showed an indication of materials with low degree of cross-linking. A Gel Permeation Chromatography (GPC) method was improved to better characterize the molecular weight distributions of the cationic structures. Preliminary qualitative cross-linking studies were performed on bisVBT-VBA copolymers, and a comparison with VBT-VBA copolymers is presented. | 1 |
99398003 | In designing VBT copolymers, one of the desirable features that previous researchers incorporated into the copolymer is the ability to be processed in aqueous systems. Due to the high level of intermolecular forces between each monomer, the VBT homopolymer is insoluble in water. Thus, VBT is often polymerized in different ratios with styrene derivative charged monomers, such as anionic vinyl phenyl sulfonate (VPS) salts [7] or cationic vinylbenzyl triethylammonium chloride (VBA) [8][9][10] which allows the copolymer to be soluble in water. Extending the knowledge achieved from the study of the natural phenomena related to thymine reactivity and intermolecular interactions has generated a diapason of potential applications [11][12][13][14] ranging from photoresist material [15,16] and organic-inorganic hybrid devices [2] to antibacterial coatings [17], drug delivery systems [3,[18][19][20], recyclable plastics [21], and biosensors [22,23]. On the other hand, the increasing attention given to the environmental and toxicological dilemmas linked to commercial materials situates the thymine-based polymeric systems as a Green Chemistry platform [24]. From an intellectual standpoint, thymine polymeric systems offer an approach to physiologically relevant processes, while offering commercially applicable opportunities in material sciences. This synergy will make available new comprehensions and the development of innovative materials. Quaternary ammonium groups, such as in VBA, are known to exhibit antimicrobial properties as they interact with the cell walls of certain types of bacteria and cause the destruction of cells by lysins. This property in 2 Journal of Polymers combination with the photo-induced immobilization of VBT monomers creates a host of possible applications for VBT-VBA copolymers, such as antimicrobial coatings in hospitals or antifouling agents on the exterior of boats and ships. Furthermore, the ethyl alkyl arms on VBA can be replaced with alkyl groups of different chain lengths to vary the antimicrobial properties and applications [17]. To ensure that the presence of charged monomers would not interfere with the photo-cross-linking reaction of VBT moieties, irradiation studies have been performed on varying ratios of VBT-VBA copolymers to quantify the extent of their photodimerization and immobilization [8,10]. From such studies, it was found that linear VBT-VBA copolymer chains cross-link through adjacent thymine moieties to form an immobilized thin film. Inspired by these findings and in the attempt to incorporate Green Chemistry [24] in our practices, we focused our research on improving the synthesis of the VBT monomer by isolating and characterizing each product/by-product. One of the most common routes of the VBT synthesis is via a nucleophilic substitution reaction between vinylbenzyl chloride (VBC) and potassium thyminate (KThy) [5]. Although this method of synthesis has been predominantly used to produce VBT, its yield has not been known to exceed 40%. Our meticulous studies permitted obtaining a high yield of VBT monomer (37.4%) and correspondingly other reaction byproducts such as 1,3-bis(4-vinylbenzyl)thymine (bisVBT) (22.1%) and trace amounts of 3-(4-vinylbenzyl)thymine (3-VBT) and 4-vinylbenzyl alcohol (VBOH). Out of these byproducts, bisVBT occurred in the highest quantity, making it the easiest to isolate and characterize. Due to the presence of only one vinyl group in its structure, the VBT monomer is only able to form linear copolymers. However, since the bisVBT monomer has two vinyl groups, it has the potential to form branched and even cross-linked networks. With the bisVBT enhanced polymerizability, it can be anticipated that the strength and flexibility of cross-linked bisVBT materials will meet the requirements of new desired applications. Furthermore, the bisVBT monomer similarly has photo-cross-linking capabilities due to the thymine base in its structure. This work describes the synthesis of three different bisVBT-VBA materials of varying molecular weights and structure and a comparison of their thermal profiles and cross-linking efficiencies to previously synthesized VBT-VBA copolymers. A Gel Permeation Chromatography (GPC) method was modified to better characterize the molecular weight distributions of the synthesized bisVBT-VBA cationic networks. Preliminary qualitative cross-linking studies were performed on bisVBT-VBA materials and a comparison with wellknown VBT-VBA copolymers is presented. PET film (Melinex 454/500) was provided by DuPont Teijin Films. Film coating was performed using a microscope slide edge to draw down the copolymer solution. Irradiation of the copolymers films was carried out using Spectronics Corp. Spectrolinker XL-1500 (254 nm, 40 W). All synthesized monomers and copolymers were dried in a vacuum oven (VWR5 1410 = 80 ∘ C; pressure = 28 inHg). Purification of VBT and bisVBT were performed using Teledyne Isco CombiFlash Rf 200. 1 H NMR spectra of monomers and copolymers were performed using a 90 MHz (Anasazi EFT) and a 400 MHz (JEOL) NMR spectrometers. IR analysis was performed using Thermo Scientific Nicolet 6700 FTIR instrument. Agilent 8453 Diode Array UV-Vis Spectrophotometer was used for sample measurement. Molecular weights were determined by GPC performed using a Beckman Gold Programmable Solvent Module HPLC system (126) with a UV/Vis Programmable Detector Module (166). A guard column and two analytical columns (Jordi DVB Polar Pack Wax mixed bed) were used. Copolymer solutions in the solvent were agitated on an orbital shaker overnight (recommended by the GPC-columns manufacturer) and filtered through a 0.45 m Whatman filter attached to a BD 1 mL Luer-Lok tip syringe before sample injection. GPC mobile phase filtration apparatus was purchased from Quark Glass. Thermal studies of the copolymers were performed as follows. TGA data were collected using a TA Instrument Q5000 TG Analyzer. In a typical TGA experiment, the copolymer sample was removed from the vacuum oven, carefully weighed, put into the furnace of the instrument, and heated, under nitrogen, over a range of 20 ∘ C-600 ∘ C on a ramp of 20 ∘ C/min. DSC data were collected using a TA Instruments Q2000 DS Calorimeter. Samples for the DSC experiment were carefully weighed, heated in Tzero aluminum hermetic pans purchased from TA Instruments. Heat-cool-heat DSC experiments were performed under N 2 flux, a heating ramp of 10 ∘ C/min, and a cooling ramp of 5 ∘ C/min. | 2 |
99398003 | Synthesis of Sodium Thyminate. Sodium hydroxide (2.04 g, 0.051 mol) was added to a flask containing DI water (30 mL), heated to 50 ∘ C, and stirred at 300 RPM until dissolved. Then thymine (6.50 g, 0.044 mol) was added and stirred for additional 20 minutes. After the solution has completely turned clear, the flask was removed from heat and cooled to room temperature. 50 mL of ethanol was added slowly to the stirring solution and a white precipitate was observed. The white solid was filtered with a Buchner funnel, rinsed with additional ethanol, and dried overnight in a vacuum oven at 80 ∘ C under 25 in Hg vacuum. Sodium | 3 |
99398003 | Synthesis and Purification of Monomers, VBT, and bisVBT. VBT synthesis was slightly modified from procedures described previously [5,6]. The reaction scheme is presented in Figure 1. In a 250 mL round bottom flask containing a magnetic stirring bar, dimethylformamide (DMF) (20 mL) was added and heated in an oil bath until the temperature was stabilized at 70 ∘ C. To the same flask, sodium thyminate (NaThy, 3.5 g, 0.024 mol) was added. Once a majority of NaThy was dissolved to give a white milky solution, vinylbenzyl chloride (VBC) (4.8 g, 0.028 mol) and inhibitor 2,6-di-tertbutyl-4methylphenol (BHT) (5 mg, 0.02 mmol) were added. The light yellow opaque mixture was allowed to heat at 70 ∘ C and stir for 24 hours. The DMF solvent was removed at the end of the reaction via rotary evaporation under vacuum. The paste-like residue was redissolved in methylene chloride and adsorbed onto silica gel. RediSep Rf prepacked silicagel columns (silica 80 g) were used to separate the reaction mixture on a Teledyne Isco CombiFlash instrument. The elution solvent system used was a gradual ramp from 20% ethyl acetate in hexanes to 50% at 60 mL/min, with a total separation time of 60 minutes. The isolated VBT (2.14 g, 37.4% yield) was a white solid. The isolated bisVBT (1.87 g, 22.1% yield) was a translucent white viscous liquid, which solidified into an opaque white solid upon cooling to 4 ∘ C. Both of the isolated VBT and bisVBT products were placed in a vacuum oven to dry and then characterized using NMR (JEOL 400 MHz) and IR. Based on 1 H NMR spectra results, the monomers were deemed pure enough for the synthesis of the copolymers. To examine the crystal packing behavior of the synthesized bisVBT monomer, crystalline samples were subjected to crystal and molecular structure determinations by Single-Crystal X-ray Diffraction. The data were collected using an APEXII BRUKER X-ray diffractometer with X-ray radiation generated from a Mo sealed X-ray tube ( = 0.70173Å with a potential of 40 kV and a current of 40 mA) at the Department of Chemistry, Texas A&M University. Detailed X-ray diffraction data are available in the Electronic Supplementary Information. Vinylbenzyl triethylammonium chloride (VBA) monomer was synthesized as described before [25]. In a threeneck 500 mL round bottom flask equipped with a stirring bar, reflux condenser, and nitrogen inlet, bisVBT (0.50 g, 1.4 mmol, 1 mol eq.) was dissolved in isopropanol (115 mL) at 80 ∘ C. Upon complete dissolution of bisVBT, the reaction temperature was brought down to 65 ∘ C. At this point VBA (11.31 g, 44.6 mmol, 32 mol eq.) was added swiftly to minimize water absorption. Following complete dissolution of VBA, 2,2 -azobisisobutyronitrile (AIBN, 0.125 g, 1% w/w) was added to the reaction mixture. The solution was allowed to stir (500 rpm) at 65 ∘ C for 24 hours under N 2 . The solution became notably more viscous as the reaction progressed. At the end of the 24-hour period, the reaction mixture was highly viscous, resembling a clear gel. The copolymer was isolated from the reaction mixture via precipitation in acetone. The mixture was vacuum filtered using Whatman grade 2 filter paper to collect the white copolymer precipitate. The collected copolymer was transferred to an empty beaker and placed in a vacuum oven to dry overnight to give 10.9 g of copolymer (92% yield). Primary analysis of the copolymer was performed using TLC (90/10 CHCl 3 -MeOH with 1% NH 4 OH mobile phase) to check for presence of monomer. Characterization of the dried copolymer was carried out using 1 H NMR spectroscopy in DMSO-d 6 , to verify the absence of unreacted monomers and the typical vinyl group signal at chemical shifts between 5 and 6 ppm was not observed in the spectra indicating the formation of a network structure. It should be noted that the copolymer swelled into a highly viscous gel when DMSO was added. | 4 |
99398003 | Synthesis of 1 : 32 bisVBT-VBA Copolymer (3% AIBN). The copolymerization procedure described above was repeated with the following modifications. The weight ratio of AIBN was increased to 3% (0.325 g). An overhead stirrer (400 rpm) was used instead of a magnetic stir bar to improve the uniformity of the reaction mixture. The bisVBT solids were ground up prior to being added in isopropanol to improve solubility. The product of this polymerization reaction was also very viscous and gel-like, similar to the previous reaction. One major difference in appearance was the presence of bubbles in the swollen product. The same acetone vortex was set up to crash out the copolymer. Vacuum filtration with grade 2 filter paper was used to isolate the copolymer particles from the mixture. The isolated white copolymer after drying under vacuum (9.02 g, 76% yield) appeared to be stickier and more hygroscopic than the previous copolymer. Preliminary analysis of the copolymer was performed using TLC (90/10 CHCl 3 -MeOH with 1% NH 4 OH mobile phase) to check for presence of monomer. Characterization of the dried copolymer was performed using 1 HNMR spectroscopy with DMSO-d 6 , to verify the absence of unreacted monomers and the typical vinyl group signal at chemical shifts between 5 and 6 ppm was not observed in the spectra, pointing to a network structure. This copolymer also swelled when DMSO was added but exhibited a lower observed viscosity than the 1 : 32 bisVBT-VBA (1% AIBN) copolymer. Terpolymer (1% AIBN). The polymerization procedure described in previous sections was repeated with the following modifications. VBT (1.35 g, 5.6 mmol, 4 mol eq.) was dissolved in isopropanol (115 mL) at 80 ∘ C. Following the complete dissolution of VBT, the temperature was decreased to 65 ∘ C, and bisVBT (0.50 g, 1.4 mmol, 1 mol eq) was added to the reaction flask allowing it to dissolve for 15-20 min. VBA (11.3 g, 44.6 mmol, 32 mol eq) was added to the reaction mixture. Once all the monomers were dissolved, AIBN (0.130 g, 1% w/w) was added to initiate polymerization. An overhead stirrer (400 rpm) was used [26]. | 5 |
99398003 | Synthesis of 1 : 4 : 32 bisVBT-VBT-VBA Observation of the reaction flask 18 hours later revealed that while the appearance of the flask contents was clear and gel-like; the stirring action of the overhead stirrer was sluggish, a clear indication that viscosity of this polymerization reaction was higher than in the 1 : 32 bisVBT-VBA copolymer reactions. The stirring rate was turned up to 550 rpm and an additional 100 mL of isopropanol was added to increase the fluidity of the mixture. After 24 hours, the reaction flask was removed from heat and stirring and the contents were processed similarly with the acetone vortex. While the reaction product was viscous, it still maintained the form of flowing liquid as compared to the gel-like consistency of the 1 : 32 bisVBT-VBA copolymer reaction products. A greater volume of acetone was required to be added to the reaction mixture before precipitated particles were observed. While it was possible to see that bisVBT-VBT-VBA terpolymer did crash out of solution, the particles were so fine that the suspension resembled skim milk. An initial vacuum filtration was performed using a fritted ceramic funnel with medium pore size, followed by centrifugation for 10 minutes at 5000 rpm and the suspension was allowed to settle by gravity and isolated by decanting. A sludgy layer of sticky white copolymer was observed at the bottom of the beaker, scraped out, and placed in the vacuum oven to dry. The dried copolymer (5.874 g, 45% yield) was a translucent white hard solid and was analyzed using TLC (90/10 CHCl 3 -MeOH with 1% NH 4 OH) to check for presence of monomer. Characterization of the dried copolymer was performed using 1 HNMR spectroscopy with DMSO-d 6 , showing the absence of unreacted monomers and confirming a crosslinked network. This terpolymer exhibited slight swelling when dissolved in DMSO and was much less viscous than the previous two copolymers. | 6 |
99398003 | Gel Permeation Chromatography. Molecular weights were determined by GPC experiments performed using Beckman Gold Programmable Solvent Module HPLC system (126) with a UV/Vis Programmable Detector Module (166). A guard column (Jordi DVB Polar Pack Wax mixed bed, 50 mm × 10 mm, 5 m particles) and two analytical columns (Jordi DVB Polar Pack Wax mixed bed, 250 × 10 mm, 5 m particles) in series were used. GPC analyses were performed by comparing the molecular weight of the sample against standards of known molecular weight. Therefore, the choice of the standards used was particularly important for the bisVBT-VBA charged copolymer systems. Poly(2vinylpyridine) standards (PVP) with molecular weights (Mp) of 1,820; 20,900, and 256,000 were purchased from Jordi Labs. The mobile phase used was 30/70 methanol/water with a total concentration of 1 M acetic acid to maintain the Journal of Polymers 5 positive charge of the column, which required a pH below 8. The solvent was made by adding 57.45 mL of glacial acetic acid to 30/70 methanol (ACS grade) and water (DI) in a 1 L volumetric flask, followed by vacuum filtration using a mobile phase filtration apparatus (Quark glass) with Nylon 66 membrane (0.45 m × 47 mm, Supelco). Copolymer sample solutions were prepared by dissolving 2.5 mg of sample per 1 mL of eluting solvent. The GPC sample solutions were agitated on an orbital shaker overnight at 150 RPM and passed through a 0.45 m Whatman filter attached to a BD 1 mL Luer-Lok tip syringe before injection. A flow rate of 1.00 mL/min was used for all runs at 254 nm wavelength. Calibration curves were obtained using poly(2-vinylpyridine) standards. | 7 |
99398003 | Irradiation Studies. A 10% (w/w) solution of each copolymer was made in 50/50 methanol and water, since it was not possible to dissolve in pure water. Upon addition to the solvent, the bisVBT-VBA (1% AIBN and 3% AIBN) copolymers exhibited a significant level of swelling, with the bisVBT-VBA (1% AIBN) copolymer swelling the most. The clear, colorless solutions had viscosities that resembled a fluid gel. The bisVBT-VBT-VBA terpolymer (1% AIBN) did not exhibit swelling and was a fluid, clear colorless solution. The PET substrate was pretreated with 1 M NaOH for 24 hours in an attempt to etch the surface and improve immobilization of the irradiated copolymer. Copolymer solutions were coated by doctor blading technique onto the PET film (Melinex 454/500) and allowed to dry for 10 min. The PET film was masked and irradiation was set for 30 sec, 60 sec, 120 sec, and 180 sec. The irradiated coatings were immersed for one minute in FD&C Blue dye solutions (0.5 g of methylene blue dye and 1 g sodium carbonate in 1 L of DI water) and then gently dipped in DI water to rinse the extra dye. | 8 |
99398003 | Results and Discussion The structure of the isolated monomer bisVBT, a by-product obtained from the VBT synthesis, was confirmed by proton NMR, IR, and X-ray crystal analysis. Moreover, three new synthesized materials were characterized by 1 HNMR and GPC. The crystal structure of bisVBT monomer and the unit cell are shown in Figures 2(a)-2(b), which reveals that the monomer molecules pack into tight layers. Nearby thymine moieties are generated by stacking of the layers, which brings the thymine groups slightly offset of the ring with an average thymine separation of 5.9Å. | 9 |
99398003 | Gel Permeation Chromatography. The average molecular weights of the resultant bisVBT-VBA copolymers were measured by gel permeation chromatography (GPC) [27]. The primary limitation of conventional GPC is that the obtained molecular weights are relative values. Therefore, the accuracy of the method depends upon the standards and samples having the same relationship between their hydrodynamic volume, charge, and molecular weight. This is especially true for charged copolymers systems [20]. Given that bisVBT-VBA copolymers are positively charged, a poly(2-vinylpyridine) (PVP) was chosen as the standard, since it becomes positively charged upon exposure to an acidic mobile condition and exhibits similar interactions with the column phase as the cationic bisVBT-VBA copolymers. A calibration curve (log Mw versus retention volume) was created from PVP standard runs, which allows determination of copolymer molecular weight based on the retention volume. Table 1 shows the weight-average molecular weight (Mw), number-average molecular weight (Mn), and polydispersity, (Pd = Mw/Mn) of the bisVBT-VBA copolymers estimated from the calibration curves. The results presented in Figure 3 show that all materials elute around 18 minutes, with average molecular weights ranging between 65,000 and 100,000 Da, and a possible secondary chain of copolymers with a slightly lower molecular weight. The differences in molecular weights for the bisVBT-VBA structures are mainly due to variations in initiator concentration [8]. The lower amount AIBN used (1 : 32 bisVBT-VBA 1% AIBN) results in longer formed chains and therefore in higher molecular weight copolymers. The fact that the terpolymer 1 : 4 : 32 bisVBT-VBT-VBA (1% AIBN) has the lowest molecular weight compared to bisVBT-VBA copolymers highlights a main difference between bisVBT and VBT monomers. The bisVBT monomer having two vinyl groups has higher quantity of sites to polymerize, giving rise to network structures and thereby increasing the molecular weight of the resulting materials as it is shown in Table 1. On the other side, the VBT monomer having only one vinyl group has lower possibilities to polymerize and generally will generate linear copolymers. Consequently, in the terpolymer 1 : 4 : 32 bisVBT-VBT-VBA with a large amount of VBT monomer present, the expected molecular weight is lower than in bisVBT-VBA copolymers. From the GPC analysis it is likely that cross-linked network copolymers from bisVBT monomers will not be soluble, and therefore the observed GPC traces of filtered samples represent the soluble fraction of the materials with low degree of cross-linking. This might explain the unexpectedly low polydispersity (Pd) values of the copolymer molecular weights, compared to reported random VBT-VBA copolymerization reactions, which exhibit acceptable polydispersity values between 1.5 and 4 [3,26,28,29]. | 10 |
99398003 | Thermal Analysis. The thermal properties and stability of the bisVBT-VBA copolymer were studied with TGA in a nitrogen stream and with DSC. The TGA thermogravimetric curves of different bisVBT-VBA copolymers and the 1 : 8 VBT-VBA control copolymer are presented in Figure 4. For all copolymers, the initial 5 to 10% weight loss at 100 ∘ C occurs as a result of water evaporation. Figure 4(a) shows that both 1 : 32 bisVBT-VBA copolymers with 1% and 3% AIBN initiator have similar features, indicating that the small difference in MW has no influence on the thermal properties. The thermograms have two very well-defined degradation steps. The first step of 40% weight loss occurring at 209 ∘ C corresponds to VBA monomer. The second degradation point for both 1 : 32 bisVBT-VBA materials corresponds to bisVBT monomer and was found at 445 ∘ C (Figure 4(a)). This value is slightly higher than the VBT degradation presented for the 1 : 8 VBT-VBA copolymer (390 ∘ C) and the 1 : 4 : 32 bisVBT-VBT-VBA terpolymer (400 ∘ C) (as shown in Figure 4(b)). While the VBA monomer ratio decreases going from 1 : 32 bisVBT-VBA to 1 : 8 VBT-VBA copolymers, the first degradation step related to VBA monomer takes place at a lower temperature, as it can be observed for 1 : 8 VBT-VBA (1% AIBN) copolymer at 195 ∘ C (Figure 4(b)). These observations suggest that bisVBT-VBA copolymers have greater thermal flexibility than VBT-VBA copolymers. An important information when evaluating new materials for their potential application is the glass transition temperature (Tg), which is defined as a change in heat capacity when the polymer matrix changes from "glass" at low temperatures to "rubber" at higher temperatures. This second-order endothermic transition appears in the thermogram as a step not a peak. From DSC thermograms (not shown) no distinguishable thermal phase transitions were observed, besides slight steps from where the Tg for each material was determined. The DSC thermograms showed a decrease in Tg values when the VBA ratio in the copolymer increases going from 89.3 ∘ C for 1 : 8 VBT-VBA copolymers to approximately 74.5 ∘ C for 1 : 32 bisVBT-VBA copolymers. This can be explained since a higher content of cationic VBA monomers increases electrostatic repulsions between monomers, which makes the material less rigid. When comparing materials having similar VBT : VBA ratio, it can be observed that for 1 : 8 VBT-VBA copolymers the Tg value (89.3 ∘ C) is higher than for the 1 : 4 : 32 bisVBT-VBT-VBA terpolymer (76 ∘ C) indicating that the presence of bisVBT monomer may act as a "plasticizer," which improves the fluidity and reduces the brittleness of the material. In general, the presence of high Tg values was expected considering that the synthesized structures have high molecular weights. The glass transition temperatures (Tg) of the bisVBT-VBA structures are lower than those of the VBT-VBA copolymers, supporting the previous conclusion that arrangements containing the bisVBT monomer have greater thermal flexibility than those containing the VBT monomer. | 11 |
99398003 | Qualitative Irradiation Studies. All three bisVBT-VBA synthesized materials and the control 1 : 8 VBT-VBA copolymer were layered on thin films and irradiated for different times 30 sec, 60 sec, 120 sec, and 180 sec (equivalent to the amount of energy delivered). The VBT films are colorless and in order to quantify the extent of their photoimmobilization on the substrate, the films were toned with an anionic FD&C Blue dye. The dye molecules have low affinity to the PET substrate, nevertheless strongly interacting with the immobilized oppositely charged polycationic copolymer. Thus, the thymine cross-linking can be indirectly estimated by analyzing the dye adsorption onto the insoluble copolymer. The immobilization degree between the substrate and the copolymer is affected by the molar mass and the chemical composition. At shorter times the photo cross-linking and the resulting increase in average molecular weight of the copolymers do not affect solubility. Nevertheless, as the reaction proceeds and more dimers are generated, the copolymer chains form high molecular weight networks, which make up the insoluble fraction of the photo-cross-linked copolymer. When all copolymer chains are connected a saturation level is reached, and continued irradiation does not cause any additional decrease in solubility. This is a typical behavior of photoresist materials [30]. Figure 5 shows qualitatively how the extent of immobilization varies between all bisVBT-VBA cross-linked materials. In all cases it was observed that the amount of trapped dye increases with irradiation time until it reaches saturation (time evolution not shown). Out of the three new synthetic copolymers, the 1 : 4 : 32 bisVBT-VBT-VBA (1% AIBN) terpolymer in 50/50 MeOH-water ( Figure 5(a)) exhibited a satisfactory degree of cross-linking, as all the photo-masked area remained immobilized. As a comparison, the previously studied copolymer 1 : 8 VBT-VBA (1% AIBN) [31][32][33] in 50/50 MeOH-water was used as a control ( Figure 5(b)). Visually, the photo cross-linking exhibited by both copolymers was comparable, as it was expected since they have similar VBT-VBA ratios. A moderate degree of cross-linking occurred with the 1 : 32 bisVBT-VBA (3% AIBN) copolymer in 50/50 MeOHwater ( Figure 5(c)), where it can be observed that longer irradiation times (120 sec) were needed to immobilize copolymer. These circumstances were expected for the reason that, when less fraction of cross-linkable VBT compared with VBA monomer is present in the copolymer films, few thymine groups undergo photodimerization, and therefore longer irradiation time is required for immobilization. Therefore, decreasing bisVBT/VBT concentration will result in thymine molecules further away from each other, and in consequence a slower and less efficient photo cross-link. In summary, copolymers with high VBA content will possess a decreased ability to cross-link when irradiated with UV light mostly due to a low content of VBT monomers, but correspondingly as a result of increased electrostatic repulsions between VBA monomers (low Tg) which hinders the free spatial arrangement of the copolymer reducing the possibility of adjacent thymine moieties to be close enough to cross-link. Figure 5(c) also shows that bisVBT-VBA structures have a typical photoresist behavior, since after 120 sec of irradiation the dye adsorption reaches saturation. While the intention of increasing the molar equivalence of VBA to bisVBT was to increase the solubility of the bisVBT-VBA materials, the smaller molar equivalence of bisVBT may have caused the copolymer chains to have insufficient amounts of neighboring thymine moieties in the proximity, required for photodimerization and adequate immobilization. Quantitative studies of the photo cross-linking of copolymers varying bisVBT to VBA ratios are currently under progress. It is worthy to note that the bisVBT-VBA copolymers in a 10% w/w solution in 50/50 MeOH-water exhibited a tendency to swell upon solvation in hydroxyl containing solvents, like water and alcohols. This property is consistent with crosslinked copolymers and may reduce the processability of the new materials, increasing the difficulty of creating uniform thickness films. | 12 |
238770759 | : In this study, the effects of an agro-industrial residue with active properties, pomegranate peel extract (PPE), were evaluated on the rheological properties of potential coatings based on chitosan (C) and gelatin (G). For this, rheological properties of the polymeric solutions were investigated in relation to PPE concentration (2 or 4 mg PPE g − 1 solution), and to its incorporation order into the system (in C or in CG mixture). All solutions were more viscous than elastic (G (cid:48)(cid:48) > G (cid:48) ), and the change in PPE concentration had a greater influence accentuating the viscous character of the samples in which PPE was added to the CG mixture (CGPPE2 and CGPPE4). PPE addition to the CG mixture increased the angular frequency at the moduli crossover, indicating the formation of a more resistant polymeric network. This tendency was also observed in flow results, in which PPE addition decreased the pseudoplastic behavior of the solutions, due to a greater cross-linking between the polymers and the phenolic compounds. In general, all the studied solutions showed viscosities suitable for the proposed application, and it was possible to state the importance of standardizing the addition order of the components during the preparation of a coating. at the moduli crossover, indicating the formation of a more intricate polymeric network. Temperature also affected the viscoelastic behavior of the samples, with CGPPE4 being the only solution with a different profile of increasing the moduli at temperatures above 65 ◦ C. Finally, the flow tests evidenced the influence of PPE in decreasing the pseudoplastic behavior of the polymeric solutions, making them more resistant to the applied shear. The results of this study contribute to the in-depth understanding of the molecular relationships between a polysaccharide, a protein, and phenolic compounds in the prepared coating solutions, and how these relationships can be related to the intended application for the materials. | 1 |
238770759 | Introduction The search for edible food coatings that are biodegradable, non-toxic, and that have active properties has been increasing in recent years, aiming to extend the shelf life of foods while preserving their sensory characteristics and improving consumer safety [1,2]. In the literature, there is an increasing number of studies that evaluate the most diverse polymers, natural or synthetic, and their physical-chemical, structural, and active properties in foods. In these studies, polysaccharides stand out as one of the groups of polymers that are most applied for coating purposes, due to their attractive properties of brightness, transparency, flexibility, barrier to the passage of water and gases, antimicrobial activity, among others [3]. Chitosan is a polysaccharide derived from the partial deacetylation of the chitin found in the exoskeleton of crustaceans and insects, as well as in the structure of mollusks and fungi [4]. Its cationicity, biodegradability, non-toxicity, and its filmogenic and antimicrobial properties make chitosan one of the most studied polysaccharides for the development of edible coatings [5,6]. To improve the mechanical and barrier properties of chitosan-based coatings, the association of this polysaccharide with other natural polymers, such as gelatin, is commonly evaluated [7]. Obtained by the partial hydrolysis of collagen, this protein is well-used in the food industry as a thickening and emulsifying agent, and several studies already successfully applied a combination of chitosan and gelatin for edible coating development [8][9][10][11]. Furthermore, studies on food coatings aim to develop materials with active properties, which can also protect the food to be coated from oxidation by light, or which | 2 |
238770759 | Materials All solvents and reagents used were PA grade and used as such. Chitosan was obtained from the partial deacetylation of β-chitin, found in the squid pens of Doryteuthis spp., obtained from Miami Comércio e Exportação de Pescados Ltd.a in Cananéia city-Sao Paulo State, Brazil. Gelatin was commercial (Sigma-Aldrich ® , St. Louis, MO, USA), type A, swine, with approximately 300 bloom. The red pomegranates (Punica granatum L.) used (Peruvian variety) were purchased at the Supply Centers of Campinas-S.A (CEASA, Campinas city-SP, Brazil) in February, 2019. Before extraction, the fruits were peeled, and their peels were sanitized with 2% (v/v) sodium hypochlorite, dried at room temperature under air flow for 2 h, frozen in the freezer and lyophilized for 16 h (Edwards equipment, model Freeze Dryer Modulyo, Burgess Hill, West Sussex, United Kingdom). Finally, they were crushed to obtain a thin powder, which was stored in polypropylene flasks under refrigeration and protected from light until use. The time between extraction and experiments was not longer than one month. | 3 |
238770759 | Chitosan Obtention and Characterization Chitosan was obtained by the partial deacetylation process, to which the β-chitin extracted from squid pens was submitted, a procedure described by Horn et al. (2009), which involves the conversion of the acetoamide groups from the (1,4)-N-acetyl-D-glucos-2-amine chains of β-chitin to amino groups, in a basic medium (40% NaOH (v/v), constant stirring for 3 h at 80 • C, under N 2 flow) [15]. The obtained powder was characterized by its viscosimetric molar mass, being classified as chitosan of high molar mass (300 kDa); its degree of acetylation of 11.1% was determined by nuclear magnetic resonance spectroscopy ( 1 H NMR) [16,17]. | 4 |
238770759 | Pomegranate Peel Extraction and Characterization The pomegranate peel extract (PPE) was obtained according to the procedure described by Bertolo et al. (2020) [9]. The powder obtained after crushing the dried peels was added to a 60% (v/v) hydroethanolic solution, in the proportion of 1 g of pomegranate peel powder to 30 g of hydroethanolic solution. The extraction took place at 50 • C, for 1 h, under slow and constant stirring. Then, the solution was filtered through a filter paper (Whatman, n. 1), and the ethanol was removed slowly by evaporation. Finally, the extract was lyophilized to obtain a dry powder. PPE was characterized in terms of its total phenolic content. For this, the Folin-Ciocalteu colorimetric method was adopted, with a procedure adapted for a 96-well microplate [18]. Gallic acid (Sigma-Aldrich ® , St. Louis, MO, USA) was used as the standard molecule for the construction of the method's calibration curve, from eight aqueous solutions with concentrations of 4,8,12,16,20,24,28, and 32 µg mL −1 . In the microplate wells, 50 µL of PPE solution (0.1 mg mL −1 ) and 50 µL of Folin's reagent were placed. After stirring and 5 min of reaction, 200 µL of a 20% sodium carbonate solution (w/w) was added to each well, followed by further stirring. All samples were taken in triplicate. After 15 min, the absorbance of the samples was read at 725 nm on a Thermo Scientific ™ UV-Vis spectrophotometer VL0L00D0 (Waltham, MA, USA). The 60% (v/v) hydroethanolic solution was used as the blank extract. | 5 |
238770759 | Coating Preparation The 1% (w/w) initial solutions of chitosan (C) and gelatin (G) were prepared as follows: chitosan was dissolved in a 1% lactic acid solution (w/w), by slow and constant stirring for 24 h; gelatin was dissolved in water at 60 • C for 30 min, and its gelation was carried out under refrigeration for 2 h. The CG control sample (that is, without PPE) was prepared by mixing the polymers in the proportion of 1:1 at 45 • C for 2 h, with the addition of 1 mL of a 50% hydroethanolic solution (v/v), in order to maintain the concentration in relation to the other samples to be prepared. Then, the incorporation of PPE into chitosan/gelatin solutions took place. For this, 2 different concentrations of a hydroethanolic solution (50%, v/v) of extract, 100 and 200 mg mL −1 , were prepared. To incorporate these solutions, two additional orders were tested: (1) Addition of 1 mL of PPE to C, followed by the mixture of G (CPPE2G and CPPE4G samples); (2) Mixture of C and G, followed by the addition of 1 mL of PPE (CGPPE2 and CGPPE4 samples). The PPE addition order to the polymeric system was changed to evaluate how the phenolic hydroxyls of the PPE components would interact with the protonated amino groups (NH 3 + ) of chitosan, in the presence (CG mixture) or in the absence (C solution) of gelatin. This protein has carboxylic groups of its amino acids responsible for the formation of the polymeric network by electrostatic interaction with chitosan (please see the GA of our previous work [7]). Thus, different orders of PPE addition to the system can lead to different rheological responses, due to the greater or lesser degree of interaction between the polymers, as well as between the polymers and the extract. Regardless of the PPE order of addition, the proportion adopted was that of 1 mL of extract for each 50 g of mixture of chitosan and gelatin. Thus, the final concentrations of PPE in the polymeric mixture were 2 mg PPE g −1 mixture (for CPPE2G and CGPPE2 samples) and 4 mg PPE g −1 mixture (for CPPE4G and CGPPE4 samples). All the prepared solutions (the controls C, G, and CG, as well as those containing pomegranate peel extract, PPE) presented a homogeneous aspect, without the presence of precipitates, and with varied yellow coloration, according to the extract concentration and its order of addition to the system. | 6 |
238770759 | Rheological Measurements The rheological study was carried out with the samples CPPE2G, CPPE4G, CGPPE2, CGPPE4, CG, and with the 1% chitosan solution (C). An AR-1000 N controlled stress rheometer (TA Instruments, New Castle, DE, USA) was used, with a cone/plate geometry of stainless-steel of 20 mm in diameter, at 2 • angle, and a fixed gap of 69 µm. The temperature was controlled with a Peltier system, and all the measurements described below were performed in triplicate. For all measurements (strain, frequency, temperature sweep, and steady shear), the prepared solutions (in the final state of a gel) were stored under refrigeration in the dark until analysis. After stabilizing the temperature of the solutions to room temperature, they were placed on the Peltier plate of the rheometer, and their excess was removed after setting the zero gap of the equipment. | 7 |
238770759 | Strain Sweep Measurements Fixed values of temperature (25 • C) and frequency (1.0 Hz) were adopted in an amplitude range of 0.05 to 500 Pa, to determine the linear viscoelastic region (LVR) of the coating solutions, that is, the strain range in which their elastic (G ) and viscous (G ) moduli do not change. The software Rheology Advantage Data Analysis, version V5.7.0 (TA Instruments Ltd., New Castle, DE, USA) was employed to analyze the parameters obtained from strain sweep measurements, which were: G LVR , the elastic modulus at the end of LVR; γ L , the maximum strain value to which the solution can be submitted before their moduli start to decrease; tanδ, the ratio between G and G ; and G -G , the difference between the moduli at 10% of strain. | 8 |
238770759 | Frequency Sweep Measurements For frequency sweep measurements, a frequency range from 0.1 to 100 rad s −1 was adopted, at fixed values of temperature (25 • C) and strain (10% in the LVR). The behavior of G and G , according to the increase in frequency, was analyzed in terms of G crossover and ω crossover , that is, G and frequency values where G = G . | 9 |
238770759 | Steady Shear Measurements Finally, flow measurements were conducted at 25 • C, with a range and shear rate of 0.1 to 1000 s −1 . The viscosity behavior of the solutions was characterized as Newtonian or pseudoplastic, according to PPE addition concentration. The experimental curves of viscosity were fitted according to the Cross model (Equation (1). In Equation (1), η 0 is the zero-shear viscosity (Pa s), η∞ is the viscosity limit at infinite shear (Pa s), γ is the shear rate (s −1 ), k is the so-called consistency index (s), and n the rate index (dimensionless). | 11 |
238770759 | Statistical Analysis The Software Action (Estatcamp Team, 2014,São Carlos-SP, Brazil) was used for the statistical analysis of the data. The Shapiro-Wilk test was applied to verify data distribution. Parametric rheological results were examined using analysis of variance (ANOVA), followed by Tukey's test. Non-parametric rheological results were examined using Kruskal-Wallis test. Significance level was set at equal or higher than 5% in all cases. | 12 |
238770759 | Pomegranate Peel Extraction and Characterization After the extraction process to which the pomegranate peels were subjected, and the lyophilization of the extract, a thin, dry, and yellowish powder was obtained, with a yield of 54.5%. Its total phenolic content (TPC), determined by the Folin-Ciocalteu method, was 213 ± 6 mg gallic acid equivalent (GAE) g −1 extract. This result is consistent with that found by Derakshan et al. (2018), who analyzed the TPC of extracts from the peel and the seed of three different types of pomegranates, finding a range of 276-413 mg EAG g −1 for the peel extracts [19]. Bertolo et al. (2020), in a previous study, determined 492 ± 82 mg GAE g −1 extract, using a yellow pomegranate of Brazilian variety [9]. It is worth mentioning that the variations observed in the TPC of different phenolic extracts are related to the most diverse factors: from intrinsic ones, such as the origin of the fruit and its harvest time, to extrinsic factors, such as the method of extraction adopted, the time of extraction, and the solvents used. | 13 |
238770759 | Rheological Measurements Strain Sweep Measurements Initially, the sweep tests of the elastic (G ) and viscous (G ) moduli of the material were performed as a function of the percentage of deformation, essential for determining the linear viscoelastic region of the solutions (LVR) where loss and storage moduli (G and G , respectively) are practically constant, regardless of the applied deformation. [14] The extent of the LVR can be directly related to the structural strength of the material: solutions that are more resistant to the applied deformation tend to have a more extensive LVR, requiring greater values of deformation for the moduli to cease stability and start to decrease [7,14]. As can be seen in the graphs of Figure 1, divided into A and B according to the order of addition of PPE in the polymeric solutions, all samples showed G > G , which indicates that their viscous behavior exceeded the elastic behavior, regardless of the addition of the extract or its concentration, a typical liquid-like behavior [7,9,10,20]. From the curves in Figure 1, it was possible to determine the parameters presented in Table 1: the first, γ L , represents the highest strain value to which the solution could be subjected, before leaving the LVR; the higher this value, the greater the resistance offered by the sample to the applied deformation. It can be said that the addition of gelatin to sample C led to a significant decrease in γ L , from 48.87 ± 3.24% in C to 40.78 ± 0.32% in CG, an indication that the incorporation of gelatin and its interaction with the polymer chains of chitosan can make the polymeric system less resistant to the applied deformation. However, the addition of PPE in CGPPE2 and CGPPE4 solutions led to an increase in their critical deformation, to values significantly equal to the original chitosan solution, not influenced by the concentration of the extract. Similar results were found by Bertolo et al. (2020) when analyzing chitosan and gelatin systems incorporated with grape seed extracts. The addition of gelatin led to a decay of the critical deformation of chitosan, and the concentration of the added phenolics changed this parameter significantly. Lower concentrations promoted greater stability for the solutions [7]. Finally, for the samples in which PPE was initially added to C, the incorporation of the extract did not significantly alter γ L , and the subsequent addition of gelatin did not promote the effect observed in the other samples, by decreasing the critical strain. However, the addition of PPE in CGPPE2 and CGPPE4 solutions led to an increase in their critical deformation, to values significantly equal to the original chitosan solution, not influenced by the concentration of the extract. Similar results were found by Bertolo et al. (2020) when analyzing chitosan and gelatin systems incorporated with grape seed extracts. The addition of gelatin led to a decay of the critical deformation of chitosan, and the concentration of the added phenolics changed this parameter significantly. Lower concentrations promoted greater stability for the solutions [7]. Finally, for the samples in which PPE was initially added to C, the incorporation of the extract did not significantly alter γL, and the subsequent addition of gelatin did not promote the effect observed in the other samples, by decreasing the critical strain. Even though γL did not undergo significant changes with the incorporation of PPE into the system, the second parameter in Table 1, G'LVR, showed more pronounced variations: the value of the elastic modulus of the solutions at the limit of LVR suffered an abrupt decrease from 22.80 ± 1.49 Pa to 4.27 ± 0.45 Pa from C to CG. The decay trend continued with the incorporation of PPE into the system, regardless of its concentration or order of addition. The third parameter in Table 1, tanδ, is the ratio between the viscous and elastic moduli at the LVR limit, and allows particularly important rheological classifications of the material: if tanδ > 1, G'' > G' and the samples are classified as viscous; the opposite is also valid, and if tanδ < 1, the elastic character predominates. However, if tanδ > 0.1, the behavior of the samples is situated between that of a highly concentrated polymeric solution and that of a real gel [20]. In the samples of this study, tanδ varied between Even though γ L did not undergo significant changes with the incorporation of PPE into the system, the second parameter in Table 1, G LVR , showed more pronounced variations: the value of the elastic modulus of the solutions at the limit of LVR suffered an abrupt decrease from 22.80 ± 1.49 Pa to 4.27 ± 0.45 Pa from C to CG. The decay trend continued with the incorporation of PPE into the system, regardless of its concentration or order of addition. The third parameter in Table 1, tanδ, is the ratio between the viscous and elastic moduli at the LVR limit, and allows particularly important rheological classifications of the material: if tanδ > 1, G > G and the samples are classified as viscous; the opposite is also valid, and if tanδ < 1, the elastic character predominates. However, if tanδ > 0.1, the behavior of the samples is situated between that of a highly concentrated polymeric solution and that of a real gel [20]. In the samples of this study, tanδ varied between 1.32 ± 0.02 (C) and 2.33 ± 0.06 (CGPPE4), which classifies them as being highly concentrated polymeric solutions, with a predominant viscous character, as had already been observed in the curves of Figure 1. The incorporation of gelatin in CG increased tanδ value from 1.32 ± 0.02 to 1.68 ± 0.06, reflecting the decrease observed in G LVR for that same sample; the incorporation of PPE into CG mixture led to values greater than two for tanδ, indicating that the incorporation of the extract accentuated the viscous character of the solutions; when doubling the PPE concentration from CGPPE2 to CGPPE4, tanδ increased significantly from 2.05 ± 0.11 to 2.33 ± 0.06. The same effect of the incorporation of PPE is valid for the samples CPPE2G and CPPE4G, with tanδ values also greater than two, but without significant changes observed when the concentration of the extract was doubled. Despite the proportion between the polymers prepared by Bertolo et al. (2020) being different (2:1), they obtained similar tanδ results for their solutions, ranging from 1.25 ± 0.16 for the sample without PPE, to 2.59 ± 0.12 for the sample with one of the highest concentrations of PPE [9]. Their results agree with the results of this study, reinforcing the tendency of phenolic compounds to increase the viscous character of the solutions. Finally, to confirm the effects of the incorporation and concentration of PPE in the polymeric system, the difference between G and G at 10% of deformation was calculated (G -G parameter from Table 1); this difference indicates whether the moduli are approaching or moving away, according to the incorporation of the phenolics, and is a new indication of which character is more pronounced. The incorporation of PPE into the CG mixture led to a significant increase in G -G difference from 2.94 ± 0.06 Pa to 3.74 ± 0.07 Pa in CGPPE2, and doubling the concentration in CGPPE4, the difference between the moduli increased to 4.04 ± 0.14 Pa; this result reinforces the effect that had already been observed, that is, that the PPE addition accentuates the viscous character of the solutions, making its difference with the elastic modulus even greater. However, when the order of PPE addition was reversed, this tendency to increase the G -G difference with twice the extract concentration was no longer observed. This result also meets the other parameters analyzed for CPPE2G and CPPE4G samples, by not showing significant differences between them, and by not following the trends observed for CGPPE2 and CGPPE4. Furthermore, these samples indicate that the addition of the extract in high concentrations to chitosan, for later mixing with gelatin, may be saturating the binding sites and making the formation of the polymeric network more difficult. Thus, even if the PPE concentration in the system is doubled, the changes observed are not as significant as those obtained in the other order of addition (Table 1). | 14 |
238770759 | Frequency Sweep Measurements The behavior of G and G moduli was also evaluated in relation to the angular frequency (Figure 2A,B). In all cases, G was predominant over G during a wide sweeping range, occurring in the crossing of the moduli and their inversion in variable angular frequency values, according to the PPE concentration and addition. This fact reinforces the classification of the samples of this study as being concentrated polymeric solutions/weak gels, since G > G at the beginning and the crossover occurred within the frequency range adopted; real gels or diluted polymeric solutions would not have the same profile [12,14]. The values of G crossover and ω crossover , that is, the elastic modulus and the angular frequency at G = G , are reported in Table 2. Chitosan solution (C) was the sample with the lowest ω crossover value (9.99 ± 0.01 rad s −1 ), and the addition of gelatin promoted an increase in that value to 19.98 ± 0.01 rad s −1 in CG, reflecting the effect of gelatin in delaying the inversion between moduli, probably due to the higher number of interactions formed between the polymers in the mixture. The addition of PPE in the CG mixture further increased ω crossover values, reaching 37.04 ± 4.64 rad s −1 in CGPPE4. It is worth mentioning that half of the PPE concentration, in the inverse order of addition (CPPE2G), presented the same crossing frequency value as CGPPE4. Again, the trend observed for CPPE2G and CPPE4G was different from the other samples: in these two cases, twice the extract concentration caused a decline in the angular frequency value to 31.69 ± 0.01 rad s −1 in CPPE4G, which can be another indication that the addition of PPE to chitosan for subsequent mixing of gelatin may make the polymeric network more susceptible to changes. In relation to the G crossover values, there was a sharp decrease from 41.54 ± 2.97 Pa from C to 14.65 ± 0.78 Pa in CG; the concentration of PPE did not promote significant changes in this parameter, but the samples in which the extract was added to the CG mixture showed, in general, G crossover values slightly higher than the others. Regarding these frequency results, Sun et al. (2020) also evaluated the behavior of G and G as a function of angular frequency for solutions based on glucomannan/carboxymethyl chitosan incorporated with epigallocatechin gallate (EGCG), one of the major phenolic compounds in tea leaves. They concluded that the addition of EGCG in high concentrations increased the values of the G and G moduli at high frequencies, implying the formation of a close non-covalent entangled network between the polymers and the phenolic [2]. Hosseini et al. (2021) reached similar findings when analyzing the dynamic behavior of film forming solutions (FFSs) based on chitosan, polyvinyl alcohol, and fish gelatin, incorporated with cinnamaldehyde. While the FFSs without the phenolic compound showed a liquid-like behavior (G > G ) at low angular frequencies, the elastic behavior (solid-like) was predominant at higher frequency values; however, once cinnamaldehyde was added to the system, the high number of interactions between the oil droplets and the polymers led to an elastic behavior that was 100%prevalent throughout the entire frequency range adopted, given the formation of an elastic solid-state [3]. | 15 |
238770759 | Temperature Sweep Measurements In rheological studies involving materials with possible applications such as food coatings, temperature is one of the main factors influencing the viscoelastic behavior that should be studied. In general, it is necessary to predict the behavior of the material (whether it will be more viscous or more elastic, and in what proportion, for example) over a wide range of temperatures, which simulate possible situations of transport, storage, and even cooking of the food to be coated. The graphs in Figure 3A,B represent the variation of G and G moduli for the samples of this study. (whether it will be more viscous or more elastic, and in what proportion, for example) over a wide range of temperatures, which simulate possible situations of transport, storage, and even cooking of the food to be coated. The graphs in Figure 3A,B represent the variation of G' and G'' moduli for the samples of this study. As observed in the previous tests, G'' > G', confirming once again the predominant viscous character of the samples. With the increase in temperature, both moduli of all samples started to decrease, a typical polymeric behavior. For the chitosan solution (C), it was observed that G'' modulus decreased in a greater proportion than G', going from 26.58 Pa at 25 °C to 5.93 Pa at 75 °C (more than 20 Pa of difference); for G', this difference between the final and the initial modulus values was not greater than 17 Pa (values obtained from the curves in Figure 3). With the addition of gelatin in CG, although the initial values of both moduli were significantly lower compared to C, the observed trend did not change; the viscous modulus continued to decrease with greater intensity as the temperature rose, reaching values lower than 1 Pa at the end of the test for almost all samples, except for CGPPE4. In this case, from 65 °C onwards, a slight increase was observed for both moduli, indicating a tendency to cross at temperatures higher than 75 °C. This tendency was not observed for any other sample, regardless of the order of addition or concentration of PPE. This may be related to a greater ease of inversion for elastic behavior at high temperatures, due to the elimination of energized water molecules and As observed in the previous tests, G > G , confirming once again the predominant viscous character of the samples. With the increase in temperature, both moduli of all samples started to decrease, a typical polymeric behavior. For the chitosan solution (C), it was observed that G modulus decreased in a greater proportion than G , going from 26.58 Pa at 25 • C to 5.93 Pa at 75 • C (more than 20 Pa of difference); for G , this difference between the final and the initial modulus values was not greater than 17 Pa (values obtained from the curves in Figure 3). With the addition of gelatin in CG, although the initial values of both moduli were significantly lower compared to C, the observed trend did not change; the viscous modulus continued to decrease with greater intensity as the temperature rose, reaching values lower than 1 Pa at the end of the test for almost all samples, except for CGPPE4. In this case, from 65 • C onwards, a slight increase was observed for both moduli, indicating a tendency to cross at temperatures higher than 75 • C. This tendency was not observed for any other sample, regardless of the order of addition or concentration of PPE. This may be related to a greater ease of inversion for elastic behavior at high temperatures, due to the elimination of energized water molecules and the consequent association between polymeric molecules [7,13]. Apparently, the addition of PPE in high concentrations to the polymeric network already formed in CGPPE4 may have facilitated the elimination of water molecules at high temperatures, leading to a rearrangement of chains with a greater increasing tendency of the moduli, which could result in an inversion between them at even higher temperatures. Luciano et al. (2021) analyzed the behavior of the viscous and elastic moduli of polymeric solutions based on gelatin with different concentrations of nisin, an antibacterial peptide, as a function of temperature. Their film-forming solutions showed G > G , and the addition of nisin was able to increase the sol-gel and gel-sol transition temperatures by 4 • C, which occurred around 14-18 • C and 25-29 • C, respectively [21]. In our study, similar transitions were not observed for the solutions containing gelatin, probably due to the high concentration of chitosan and the formation of an intricate polymeric network between the polymers. In general, we can conclude the discussion of oscillatory rheology results by saying that both factors of concentration and order of addition of PPE contributed to the differences observed in the results of deformation, frequency, and temperature. The viscous and elastic moduli of the samples can be influenced to a greater or lesser extent, according to the highest or lowest concentration of extract, as well as if it was added to the polymeric network already formed or to the chitosan solution first. For CGPPE4 sample, the effects of the highest concentration of PPE were noticeably clear, leading to a sample with a higher viscous character, with inversion between the moduli in higher angular frequency values, and with a possible crossing between them in temperature values lower than the others. Figure 4A,B shows the viscosity curves of the samples as a function of the shear rate; in all cases, a decrease in viscosity was observed according to the increase in shear rate, a typical polymeric behavior known as shear-thinning or pseudoplastic. This behavior is associated with the fact that the polymeric chains orient themselves towards the applied stress, increasing their ordering and, consequently, decreasing viscosity [22]. | 16 |
238770759 | Steady Shear Measurements of the highest concentration of PPE were noticeably clear, leading to a sample with a higher viscous character, with inversion between the moduli in higher angular frequency values, and with a possible crossing between them in temperature values lower than the others. Figure 4A,B shows the viscosity curves of the samples as a function of the shear rate; in all cases, a decrease in viscosity was observed according to the increase in shear rate, a typical polymeric behavior known as shear-thinning or pseudoplastic. This behavior is associated with the fact that the polymeric chains orient themselves towards the applied stress, increasing their ordering and, consequently, decreasing viscosity [22]. Knowing the viscosity of the polymeric solution that will act as a coating is extremely important to predict its behavior. In general, we look for samples with intermediate viscosities, which escape from the extremes of high viscosity (which would be an obstacle to coat food by immersion or spray, for example) and low viscosity (which would make it Knowing the viscosity of the polymeric solution that will act as a coating is extremely important to predict its behavior. In general, we look for samples with intermediate viscosities, which escape from the extremes of high viscosity (which would be an obstacle to coat food by immersion or spray, for example) and low viscosity (which would make it difficult to spread the solution over the food surface, due to its dripping effect). The literature points to viscosities in the range of 1-10 Pa s as being suitable for experimental coating processing conditions [23]. | 17 |
238770759 | Steady Shear Measurements As can be seen in Table 3, the adjustment of the viscosity data with the Cross model allowed for the identification of some important parameters. The first one is η 0 , the initial viscosity of the solutions at zero shear rate, when the polymeric molecules are still randomly oriented, except for sample C, which had the highest initial viscosity value (η 0 = 14.90 ± 1.27 Pa·s). All samples containing gelatin and/or PPE showed viscosities between 1.92 and 2.85 Pa s, an indication that all of them are suitable for the proposed application. Although no significant differences were observed (p < 0.05), according to the addition of PPE or its concentration, some trends can be elucidated. The addition of PPE to the polymeric network already formed in CG tended to decrease its viscosity, probably due to the interaction of the extract in the active sites that had not yet been occupied by the polymers. The increase in its concentration in CGPPE4 followed the same decreasing tendency, with a viscosity of 2.05 ± 0.18 Pa s. In the samples in which PPE was initially added to chitosan, η 0 values were slightly lower than 2 Pa s, and CPPE4G showed the lowest viscosity value among all samples, of 1.92 ± 0.14 Pa s. The second parameter presented in Table 3 is k, the so-called consistency index: the lower k, the greater the Newtonian plateau of the sample before its pseudoplastic behavior begins, that is, the greater the value of the critical shear rate necessary for the sample viscosity starts to decrease [14]. Sample C was the one with the highest k value (0.212 ± 0.020 s), and the addition of gelatin in CG decreased this value to 0.119 ± 0.004 s. This indicated that the formation of the polymeric network tended to better stabilize the polymeric chains, making them most resistant to applied shear and slowing the decline in viscosity. The k index continued to decrease with the addition of PPE and the increase in its concentration, with CGPPE4 sample having the lowest k (0.056 ± 0.001 s) and, therefore, being the most resistant to shear. For CPPE2G and CPPE4G samples, twice the PPE concentration did not significantly change k, and the values remained between 0.066 and 0.069 s. Finally, the last parameter, n, the rate index, represents the dependence of viscosity in relation to the shear rate: samples with values of n between zero and one are considered pseudoplastic, while values of n greater than one characterize Newtonian samples [24]. For the samples in this study, n varied from 0.744 (CGPPE2 and CPPE2G) to 0.800 (C), which reinforces the pseudoplastic behavior of all of them, and indicates that the addition of PPE promotes a slight decline in it. The results found here for k and n parameters agree with that observed by Tudorache & Tordenave (2019). They analyzed the pseudoplastic behavior of different polysaccharides (β-glucan, xanthan gum, and guar gum) complexed with phenolic compounds (vanillin, ferulic acid, acid caffeine, among others); in all cases, the addition of phenolics also caused a decline in the pseudoplastic behavior of the samples. At a molecular level, these results can be interpreted in view of the weak associations existing between the polymeric chains, which flow easily upon shearing, presenting pseudoplastic behavior. Once the phenolic compounds are added, they mediate a stronger cross-linking between the polymeric molecules, making them more resistant to the applied shear, which explains the accentuation of the Newtonian behavior for samples containing PPE [25]. Rodrigues et al. (2020) also evaluated the rate index of solutions based on chitosan and gelatin incorporated with extracts of grape seed and jaboticaba peel; in their case, it was possible to observe a decline in the values of n (and a consequent decrease in the pseudoplastic behavior) only for the highest concentrations of extract (5 mg g −1 mixture) when added to the mixture of chitosan and gelatin already formed. In this sense, PPE was able to promote more marked changes in the behavior of the polymeric system, even in lower concentrations [10]. Thus, it can be said that the rheological results of flow agree with the oscillatory results previously discussed for the samples of chitosan and gelatin with pomegranate peel extract. It was observed that the order of addition of PPE to the polymeric matrix is a parameter to be standardized during the formulation of new coatings, since it is able to change their rheological characteristics. The concentration of PPE was again a factor of great influence, especially for the samples in which the extract was added to the polymeric network already formed by CG. | 18 |
102464998 | Five types of red wine produced from the grape varieties from the Metohia region (vintage 2014) were characterized on the basis of their chromatic properties. The properties of bottled wines: Merlot, Vranac, Prokupac, Cabernet and Game were analyzed. Chromatic characteristics of these wines were observed four times during the year – spectrophotometer measurements were performed on wines aged 0, 4, 8 and 12 months. Intensity, hue and brilliance of color of these wines were determined (by the usual method of Glories). The amount of coloring matter was determined by the usual method of Durmishidze and the percentage of polymeric anthocyanins was calculated as well. Wine ageing decreased the color intensity while the color hue value increased. It was also found that contribution of wine color red pigment decreased with wine ageing, while the percentage of yellow pigment in wine increased. The total amount of colored substances in wines studied decreased with wine ageing, while the percentage of polymeric pigments in wine increased. This study presents the methodology for analyses of the chromatic characteristics, and explains the origin influence of wine on these properties. On the basis of these correlations the quality of red wine can be established. | 1 |
102464998 | Introduction Present-day studies point to the fact that of all the foods and beverages that people consume wine is the most important source of substances playing a protective role against cardiovascular diseases, cancer and neurodegenerative diseases (Fuhrman et al., 1995;Wood et al., 1982;Okuda et al., 1992).The protective role comes from components derived from non-alcoholic wine comprising the wine polyphenols, tannins and anthocyanins in particular, which have a high antioxidant activity.Numerous studies have shown that geographical origin and grape varieties have a significant effect on the antioxidant and also on the red wine color (Budić-Leto et al., 2003;Kovač et al., 1995). Phenolic compounds are a widespread group of plant metabolites, which can be of a very simple structure, such as phenolic acids, or of a complex structure, i.e. polycondense compounds, such as proanthocyanidins (Lekuta et al., 2005). Four main groups of phenol compounds include phenolic acids, flavonoids, anthocyanins and tannins.The flavonoids are yellow-hued derivatives of flavone, where R and R1 are substituents (Figure 1).A related compound is the major red pigment in wine, malvidin-3monoglucoside, and anthocyanin (Figure 2). | 2 |
102464998 | Spectrophotometric analysis of the red wine color The spectrum of red wine has a maximum at 520 nm, due to anthocyanins and their flavylium combinations, and a minimum in the region of 420 nm.Color intensity and hue, as defined by Sudraud (1958), only take into account the contributions of red (520 nm) and yellow (420 nm) overall colors.Of course, the results of this partial analysis cannot claim to reflect the overall visual perception of a wine color.The current approach to color analysis in winemaking requires optical density measurements at 420 and 520 nm, with an additional measurement at 620 nm to include the blue component in young red wines, so called the Glories method (1984).These measurements are used to calculate the values employed to describe wine color. Wine color intensity (I) is an amount of color.It varies greatly in different types of wine: Wine hue (T) indicates the development of a color towards orange.Young wines have a value on the order of 0.5-0.7 which increases throughout aging, reaching an upper limit of around 1.2-1.3.Chromatic structure, i.e. the contribution (in percentage) for each of the three components of the total color: The brilliance of red wines (d) is associated with the shape of the spectrum.When the wine is bright red, the maximum spectrum at 520 nm is narrow and well defined.On the other hand, the maximum of the spectrum is relatively broad and flattened when wine is deep red or brick red.This feature can be presented as follows: Expected results are between 40 and 60 in young wines.A higher value indicates a dominance of red wine (Rib'ereau-Gayon et al., 2006).Spectrophotometric determination of the amount of coloring matters and the percentage of polymeric anthocyanins in red wine Coloring matters of red grapes and wine are anthocyanins.They are among the most important plant pigments.The grape and wine anthocyanins are mostly in the form of glycosides.Grape variety, V. vinifera, is characterized mainly by the presence of monoglucoside.The most common anthocyanins in grapes and wine are malvidin, peonidin and cyanidine.In the pink wine, the colored matter content ranges from 50 to 100 mg/dm 3 , in normal-colored red wines from 100 to 200 mg/dm 3 , and in highly colored wines, it can occur in up to 500 mg/dm 3 . Before the spectrophotometric measurement of the amount of colored substances in a solution is performed, it is necessary to determine the light wavelength to be used in the analysis.Experimentally, it has been found that in red wines the strongest absorbance of light of wavelengths is around 530 nm, therefore, in the spectrophotometric determination of the amount of colored substances in red wine, this light of wavelength is used (Blesić, 2006). Among the numerous methods for the spectrophotometric determination of the content of colored substances in red wine, due to the simple execution and obtaining the results of satisfactory accuracy, the method Durmishidze (1958) has been used.This method is based on the spectrophotometric determination of transparency of the defined wine solution layer and on the basis of that, it leads to the indirect determination of the content of colored compounds using Durmishidze's table, which specifies the amount of colored substances (mg in 10 cm 3 of wine, diluted according to the procedure by the author). The determination of the percentage of polymer of anthocyanin is performed by chemical treatment of the analyzed wine by sulfur dioxide, by the Russo method (2011).Sulfur dioxide, a product of sodium metabisulfite, bleaches any monomeric anthocyanins.The residual color of the wine is derived from polymerized phenolic compounds, mostly from anthocyanins.The percentage of polymer of anthocyanin is calculated as the ratio of the absorbance obtained for the wine tested at 520 nm to the absorbance for the same wine treated with sulfur dioxide: | 3 |
102464998 | Material and Methods In this study, we analyzed the red wine produced from the varieties of Metohija regions (vintage 2014).The properties of wines: Merlot, Vranac, Prokupac, Cabernet and Game were examined.Chromatic characteristics of these wines were analyzed four times during the year, and measurements were made on the wine aged 0, 4, 8 and 12 months.We considered a very young wine (zero months) the wine that was the first to be decanted and filtrated.This process enables the essential clarity of wine samples. For spectrophotometric measurements, Spectrofotometer UV-9200 RAY LEIGH was used.The experimental part was structured into the following main parts: • Analysis of the color of red wine (chromatic parameters); • Determination of the amount of coloring matters and the percentage of polymeric anthocyanins in red wine. | 4 |
102464998 | Spectrophotometric analysis of the color of red wine For this type of analysis, the Glories method was used.The spectrophotometer was equipped with the optical path length cuvettes of 1 mm, and included the possibility of reading the absorbance at light wavelengths at 420, 520 and 620 nm.The wine used for analysis must be completely clear.The intensity, hue and brilliance of tested red wines were determined spectrophotometrically by measuring the absorbance at 420, 520 and 620 nm.Distilled water was used as a blank solution. Values of parameters of intensity and hue, as well as brilliance of wines were calculated according to the forms set out in the introductory section of this paper. | 5 |
102464998 | Spectrophotometric determination of the amount of coloring matters and the percentage of polymeric anthocyanins in red wine For spectrophotometric determination of amounts of colored substances in red wine the method of Durmishidze can be used.Contents of colored matters in red wine were expressed in mg/dm 3 .Determination of the percentage of polymeric anthocyanins was performed by the Russo method. | 6 |
102464998 | Results and Discussion The data presented in Table 1 show chromatic parameters of examined wines obtained by the Glories method.By the use of this method, apart from intensity, hue and brilliance of red wine, the contributions of each pigment (yellow, red and blue) to wine color were also determined. The contribution of the red pigment was most pronounced in very young Vranac and Merlot wines (48.0% and 48.6%), while the lowest contribution was pronounced in 12-month-old Prokupac and Game wines (40.9% and 41.4%).It can be concluded that the percentage of red pigment in the color of wine increases with the aging (Birse, 2007;Harbertson and Spayd, 2006;Poiana et al., 2007).In contrast, the percentage share of the yellow pigment of wine decreased.The participation of the blue pigment in wine color was by far the least attended, it ranged from 10.6% for the young Merlot wine to a maximum of 21.2% for the 12month-old Prokupac wine.The share of the red pigment was predominant in all analyzed wines and ranges, depending on the type and age of the wine from 41.7% to 48.0%.The contribution of the yellow pigment to red wine color ranged from 36.1% to 42.3%, which is correlated with information that can be found in literature (Birse, 2007;Harbertson and Spayd, 2006;Poiana et al., 2007).The highest value of color intensity (I) was observed in young wines, especially in Merlot and Vranac (2.26 and 2.24) and the lowest value was recorded for Prokupac, aged 12 months -1.18.With wine ageing the color intensity slightly decreased.In contrast to the intensity, the value for the color hue (T) slightly increased with the process of the wine ageing (Birse, 2007;Harbertson and Spayd, 2006;Poiana et al., 2007).Thus, the maximum value for hue in a very young wine was found in Merlot -0.86, and in 12-month-old wines it is was found in Cabernet -0.99.Brilliance of wine also decreased with wine ageing and it was most pronounced in young Vranac -47.1 and the lowest value was found in 12-monthold Prokupac -27.7. With wine ageing, the value measured at λ = 520 nm decreased, which was accompanied by an increase in the measured values of the wavelengths 420 nm and 620 nm.This can be explained by transition of monomeric anthocyanins into polymeric anthocyanins (Pasku, 2005). Table 2 contains the values for the amount of colored substances in the red wines tested.With wine ageing, this value decreased, which was fully in line with the decline in the share of the red pigment in the color of wine.The highest value for the amount of colored matters was found in Merlot, in which the value of wine aging decreased from 309 mg/dm 3 to 226 mg/dm 3 .The lowest value of this parameter was recoded for Game wine, and that value of wine aging decreased from 150 mg/dm 3 to only 70 mg/dm 3 . 1M -Merlot, 2 V-Vranac, 3 P -Prokupac, 4 C -Cabernet, 5 G -Game, a X -Durm. In Table 3, the values for the percentage of polymeric anthocyanins in the wines tested are given.These values increased with wine ageing.Thus, the lowest percentage of polymeric anthocyanins was found in the young Merlot wine, 42.00%.With the ageing of this wine, that percentage increased to 62.00%, for the wine aged 12 months.The highest percentage of polymeric anthocyanins was found in the wine variety Cabernet and it ranged from 64.42% for a very young wine to 84.82% for the wine aged 12 months.From the data shown in Tables 1, 2 and 3, it can be concluded that the aging of the wine is characterized by its chromatic change in the structure -it leads to the stabilization of color.The percentage of polymeric pigments in the color of wine increased by the aging process of wine.Namely, in this process the monomeric anthocyanins turn into polymeric anthocyanins with different molecular mass.In practice, this phenomenon of color evolution of red wine is called "wine ageing".Stabilization of wine color is attributed to the reduction of participation of monomeric and copigmented anthocyanins in the wine content and the formation of combinations of tannins and anthocyanins -polymeric pigments which are characterized by red color.These polymeric pigments are highly stable compounds responsible for the color of old, red wine.Copigmented anthocyanins are complex compounds which result from the reaction between anthocyanins and copigmented molecules. | 7 |
34721866 | Horky, D.: Submicroscopic Structure of Synovial Membrane in the Adult Pig. Acta vet. Bmo 60,1991: 3-13. The synovial membrane of adult pigs was investigated. Samples were obtained from hip joints of pigs of both sexes at 15 to 24 months after birth. The tissues were processed in the routine manner to be examined by light and transmission electron microscopy. The synovial membrane in adult pigs involved two types of synovialocytes, A and B, which were arranged on its surface in 2 to 3 layers. Type A cells near the surface presented as single cells while B cells formed small clusters. These contained. apart from typical fully differentiated cell types, also transient A-B types which had all characteristics of A cells together with bodies corresponding in size and appearance to secretory granules of B cells. The cytoplasm of both A and B cells showed the presence of intracytoplasmic filaments. Type A cells had the basal membrane while in type B cells this was absent. Near the membrane surface the fibrillar component of synovial matrix consisted of collagen fibrils which, in areas penetrated with synovialocyte projections, were unmasked and protruded into the articular cavity. In surface layers, aperiodic filaments were prevailing while towards deeper layers increasing numbers of typical collagen fibrils running in various directions were observed. Aperiodic collagen fibrils penetrating through the B-cell membrane were seen repeatedly. When approaching the cell membrane they attained a periodic appearance. SynOfJial membrane, A, B, A-B synOfJialocytes, matrix synuvialis The synovial membrane plays a major role in both physiology and pathology of the joint. This has been a reason for thorough studies of its building units, i.e. cells and intercellular matter. Even though these building elements have been investigated for nearly 250 years (Hunter 1743 see Ghadially 1983), the information is still incomplete. At first attention was given to the synovial membrane of adults in experiments with mammalian animal species and later in man. Many authors have studied and described the microscopic structure of synovial membrane under physiologic, experimental and pathologic conditions (for review see Horky 1981; Ghadially 1982). The present trend, which is to gain a deeper insight into its structure and particularly into the development of its functions in relation to advancing differentiation, makes the use of young unmature animals including embryos (for review see Horky 1984, 1989ab). The submicroscopic structure, histochemical and cytochemical properties, and immunohistochemical characteristics have been reported under physiologic and pathologic conditions in different mammalian and avian species (Langer and Huth 1960; Barland et a1.1962; Cutlip and Cheville 1973; Horky et a!. 1975; Fell et al. 1976; Linek and Porte 1978; Horky 1981, 1984, 1989ab; Ghadially 1982; Okada etal. 1981; Tofft and Ef'fendy 1985; Karatzias et al. 1986; Gaines et a1. 1987; Itokazu et al. 1988; Lukoschek et al. 1988) and others. These observations revealed the presence of two types of synovial cells in the synovial membrane of all the species so far studied. These are: i) A or M (macrophage-like) cells reminiscent of histiocytes by their structure, showing phagocytic properties (Ball et a1.1964; Fell et al. 1976; Horky et al. 1974; Mapp ad Revell 1988); ii) B or F (fibroblast-like} cells or S (secretory) cells which, in | 1 |
34721866 | The synovial membrane plays a major role in both physiology and pathology of the joint.This has been a reason for thorough studies of its building units, i.e. cells and intercellular matter.Even though these building elements have been investigated for nearly 250 years (Hunter 1743 -see Ghadially 1983), the information is still incomplete.At first attention was given to the synovial membrane of adults in experiments with mammalian animal species and later in man.Many authors have studied and described the microscopic structure of synovial membrane under physiologic, experimental and pathologic conditions (for review see Horky 1981;Ghadially 1982).The present trend, which is to gain a deeper insight into its structure and particularly into the development of its functions in relation to advancing differentiation, makes the use of young unmature animals including embryos (for review see Horky 1984. The submicroscopic structure, histochemical and cytochemical properties, and immunohistochemical characteristics have been reported under physiologic and pathologic conditions in different mammalian and avian species (Langer and Huth 1960;Barland et a1.1962;Cutlip and Cheville 1973;Horky et a!. 1975;Fell et al. 1976;Linek and Porte 1978;Horky 1981Horky , 1984, 1989ab;, 1989ab;Ghadially 1982;Okada etal. 1981;Tofft and Ef'fendy 1985;Karatzias et al. 1986;Gaines et a1. 1987;Itokazu et al. 1988;Lukoschek et al. 1988) and others.These observations revealed the presence of two types of synovial cells in the synovial membrane of all the species so far studied.These are: i) A or M (macrophage-like) cells reminiscent of histiocytes by their structure, showing phagocytic properties (Ball et a1.1964;Fell et al. 1976;Horky et al. 1974; Mapp ad Revell 1988); ii) B or F (fibroblast-like} cells or S (secretory) cells which, in most mammals, are characterized by a well-developed granular endoplasmic reticulum, a large Golgi complex and the presence of secretory granules.These cells have been reported by Barland et a1. (1962), Johanson and RejnO (1976), Okada etal. (1981), Horky (1984, Graabaek (1984) and others in the whole range of mammalian species. The synovial membrane in pigs has so far received little attention.The first more detailed data were published by Roberts et al. (1969) who studied the structure of synovial membrane in femoropatellar and tibiotarsal joints in pigs at 1 day and at 2 months after birth.Further results were reported by Fell et al. (1976) who made investigations of synovialoeyte structure in metacarphophalangeal joints of pigs at 18 weeks of age and of cell behaviour in synovialocyte cultures grown in vitro for 6 days.Horky has reported (1989ab) the submicroscopic structure of synovial membrane of the hip joint in the prenatal and early postnatal periods and has also been concerned with the arrangement of intercellular matter of synovial membrane.These studies have been completed by the results presented in this paper which deals with the submicroscopic structure of synovial membrane in adult pigs. | 2 |
34721866 | Materials and Methods Samples of porcine synovial membrane were obtained from 5 pigs of both sexes at 15 to 24 months after birth.The tissue was collected in all instances from the hip articular capsule and processed for examination by light and electron microscopy.The samples of synovial membrane including part of subsynovial tissue were carefully dissected into strips (1 by 1 by 2-3 mm) in a drop of fixation liquid.Immediately, the strips were fixed in glutaraldehyde (300 mmol/l) in 0.1 M phosphate buffer (PH 7.4) for 60 then 180 min.and rinsed in three fresh baths of 0.1 M phosphate buffer (PH 7.4).Fixation was carried out in 40 mmol/l OsO, in phosphate buffer (PH 7.4) for 15 and 45 min.The specimens were dehydrated in two baths with anhydrous acetone for 30 min.each.Immersion was performed in the standard way and the tissues were embedded in Durcupan ACM.Polymerization took place in an oven at 60°C for 3 days.Ultrathin sections were cut with an Ultracut Reichert ultramicrotome, stained with lead citrate according to Reynolds or with 1 % uranyl acetate followed by lead citrate.The sections were examined and photographed with a Tesla BS 500 microscope.Seinithin sections for light microscopic observations were made from the same Durcupan embedded blocks and stained with 1 % methylene blue and Azure II. | 3 |
34721866 | Submicroscopic structure of synovial membrane cells The synovial membrane of adult pigs was covered with three or, in some regions four layers of cells.Synovialocytes were embedded in the matrix synovialis which made up a thick layer above them towards the articular cavity (Fig. 1).In the opposite direction they continued into the subsynovial tissue without forming any sharp boundary.Type A and type B cells were unevenly distributed in the respective layers of the synovial membrane. | 4 |
34721866 | Ultrastructure of type A cells A cells in the porcine synovial membrane were less numerous than B cells.They were seen near the membrane surface as single cells (Figs 1,2,3) or in aggregates of 2 to 3 cells among type B cells.Apart from typical A cells, transient A-B form were also found. | 5 |
34721866 | Nucleus It was usually oval in shape (Fig. 2) and about 8 by 3-4 pm in size.The nuclear envelope of the usual arrangement sent wide shallow invaginations against the karyoplasm (Fig. 2).The perinuclear shape was mosdy narrow, but in some areas was dilated up to 0.1-0.2pm (Fig. 2).The zonula nuc1eum limitans was seen as a continuous line (maximum 0.1 pm) along the inner membrane of the nuclear envelope (Fig. 2).Chromatin was arranged into a continuous layer at the nuclear periphery or was found as small karyoso~es on sections through the nucleus.The continuous layer was closely attached to the zonula nucleum limitans and on few occasions was interrut>ted with nuclear pores (Fig. 2). Nucleoli were rare and, when seen, of reticular types. | 6 |
34721866 | Cytoplasm The granular endoplasmic reticulum was observed in the cytoplasm of A cells as few short and flattened cisternae (Fig. 1, 2) with scarce ribosomes. The agranular endoplasmic reticulum, most pronounced in A cells near the membrane surface, formed large vacuoles and small vesicles (plate 1.), Fig. 1).The big vacuoles usually contained material of varying appearance and density.It is possible that some of the vacuoles were secondary lysosomes.The small vesicles were either coated-vesicles or more often vesicles of uniform appearance likely to have pinocytotic function.Identical structures could also be seen on sections through cytoplasmic projections of A cells (Fig. 1).Some of the 0.3-0.4p.m vesicles Plate II., (Fig. 2) were filled with finely granulated, medium electron-dense material.These vesicles were regarded as transport vacuoles. The Golgi complex was not well developed in the cytoplasm of A cells.It included occasional smooth vacuoles and the transport vacuoles mentioned above. Mitochondria had the usual structure and were few in number.Some had an elongated shape and attained a length of2-3 p.m. Apart from them, some forms with markedly dense matrix were also seen (Fig. 1).Mitochondria with transparent matrix (Fig. 2) were observed mostly in cells' cytoplasmic projections. Ribosomes were numerous and, apart from the few ribosomes attached to the outer membrane of the nuclear envelope and to cisternae of the granular ~ndoplasmic reticulum, they were diffusely distributed in the cytoplasm.This gave the cytoplasm a dark appearance. No centrioles were found in A cells of the synovial membrane of adult pigs.Because synovialocytes were not in close contact with each other, the intercellular supporting structures such as zonula occludens, ~onula adherens and desmosomes .could not be seen either. Cell membrane.Cross sections showed the C-cell cytoplasm sending many projections, several p.m long and considerably thick, into the surrounding matrix synovialis (Fig. 2).The projections contained cytoplasm with all the cell organelles and a markedly high number of pinocytotic vesicles (Fig. 1). Neither lipid droplets not glycogen were observed in the tissues under study.Intracytoplasmic filaments occurred only occasionaly as thin bands of fibres, 5 p'm thick, scattered in the cytoplasm.These were vimentine filaments belonging to the cytoskeleton structures.Type A cells were found to be separated from the surrounding matrix with a distinct, though partly incomplete, basement membrane (Fig. 1, 2). | 7 |
34721866 | Ultrastructure of transient type cells Apart from typical type A and type B cells, the synovial membrane also included transient type cells (A-B cells) which combined submicroscopic characteristics of both types.They were seen occasionally within groups of 2 to 3 type A cells present in the vicinity of B cells. | 8 |
34721866 | Nucleus It was pear-shaped and had similar dimensions as A-cell nuclei.The nuclear envelope had the usual arrangement with the exception of the perinuclear space which was distended along the whole nuclear periphery (Plate III., Fig. 3).The inner nuclear membrane was lined with zonula nucleum limitans corresponding in appearance and thickness to that of A-cell nuclei.Chromatin was aggregated into large karyosomes situated near the nuclear envelope.The rest of the nucleus was transparent containing a low number of small chromatin clusters.Occasional perichromatin granules were present (Fig. 3).The size of a compact type nucleolus was 2.5-3 pm.(Fig. 3). | 9 |
34721866 | Cytoplasm The granular endoplasmic reticulum consisted of short flattened cisternae and broadly dilated sacks with amorphous or filamentous medium electron--dense material. The agranular endoplasmic reticulum was arranged in a way typical of A cells.Near the surface there were numerous large vacuoles (1-1.5 pm) with an electron-transparent appearance.In addition, there were unevenly distributed small smooth vesicles filled with finely granulated or filamentous material and large numbers of small smooth vesicles of agranular endoplasmic reticulum. The Golgi complex was well formed.It involved several fields and its structures were usually dilated (Fig. 3).Its cisternae sequestered small Golgi vesicles with dark, either granulated or homogeneous content.Most of these remained in the vicinity of the complex but some could be seen scattered in the cytoplasm or cytoplasmic projections.They were similar in appearance to secretory granules ofB cells (see below) or to transport vacuoles of A cells. Mitochondria had the usual appearance and arrangement.No dark or very light mitochondria with damaged cristae or clear matrix were observed. Ribosomes did not differ from those of B cells in either amount or arrangement. Lysosomes were a regular finding in the cytoplasm of transient type cells.They were present in the cytoplasm near the nucleus or in cytoplasmic projections (Fig. 3). Similarly to A cells, no centrioles were found in the transient type cells.Cell membrane.The cytoplasm of transient type cells produced a few short cytoplasmic projections (Fig. 3).These contained cytoplasm of the same composition as was that found close to the nucleus.The cell membrane produced a lower number of pinocytotic vesicles than that in A cells and the basement membrane was not formed. Intracytoplasmic filaments showed no difference in either appearance or arrangement as compared with those in synovialocytes of type A cells. | 10 |
34721866 | Ultrastructure of type B cells When compared to type A cells, B cells were found to be the prevailing cell population of the synovial membrane.They were observed in the deeper parts of the membrane in the form of groups or small clusters embedded in the synovial matrix.In contrast to A cells, they took the shape of an irregular polyhedron, an irregular oval or an elongated cylinder (Plate IV., Fig. 4). | 11 |
34721866 | 'Nucleus It was irregular in shape.Its size was about 7.5 by 3.5-4 #m.The nuclear envelope arranged in the usual manner formed deep invaginations against the karyoplasm (Fig. 4) thus giving the nucleus a lobular appearance.The zonula nucleum limitans had the same width as in A cells.Chromatin was concentrated into a narrow layer along the nuclear periphery, leaving the remaining nucleus to appear very light except for several where perichromatin bodies were surrounded with chromatin.The accumulation of chromatin, presenting as so-called perinucleolar chromatin, could be seen near the nucleolus.Nuclear pores were more numerous than in the other cell types. Cross sections through the nucleus regularly revealed nucleoli, 1-1.8 #m in size (Fig. 4) which always included nucleolonema.On rare occassions segregation ()f the pars granulosa or the pars fibrosa was seen.Cytoplasm B cells had cytoplasm with a markedly large granular endoplasmic reti-'CuI um (Figs. 4,5,6).This consisted of many narrow cisternae varying in length, filled with medium-osmiophilic, finely granulated or filamentous substance (Plate V., Fig. 5, 6).Even though the endoplasmic reticulum was very rich in cisternae, no close relation to any of the other organelles was revealed.In some type B cells the cytoplasm near the nucleus had a region where membrane structures were arranged in parallel layers.These were reminiscent of anular membranes (membranae annulatae) (Plate VI., Fig. 7). The agranular endoplasmic reticulum presented as occasional smooth vesicles, 0.2-0.3#m, both in the cytoplasm and near the cell membrane.The vesicles were electron-transparent. The Golgi complex was well developed.It spread over several fields (Figs 4, 5, 6, 7, 8) taking up a large part of the cytoplasm.The structure was typical.The dilated cisternae sequestered small and large Golgi vacuoles which passed to the surrounding cytoplasm as secretory granules, lysosomes or transport vacuoles (Figs 4,5,6,7,8). Mitochondria had the usual structure.The most frequently occurring mitochondria were 1-1.5 #m long with markedly dark matrix (Figs 4, 5).Their number was much higher than in A cells. Lysosomes were hardly discernible in the electron micrographs.Since no histological proof was produced, it was not possible to distinguish them from secretory or transport granules. Secretory granules were conspicuous structures of the B-cell cytoplasm.They occurred in large numbers and attained a size of 0.5-0.8#m.They were bounded with a smooth membrane and contained granular or homogenous material of varying density (Figs 4,5,7,8).Some of them had lighter peripheries and darker centres (Plate VI., Fig. 8), which made them reminiscent of heterogenous prozymogen granules.Other granules (Fig. 7) were similar to coated vesicles. Centrioles were frequently present in the B-cell cytoplasm (Figs 4, 5).They showed the usual arrangement and were seen in various parts of the cytoplasm. Cell membrane.In type B cells the cytoplasm occasionally sent out short thin projections and thick projections.The thin ones were free from organelles ()r secretory granules, while the cytoplasm of the thick projections had the same content as that of the cell (Figs 7,8).The cell membrane segregated a lot of .pinocytotic vesicles (Figs 4,7). Intracytoplasmic filaments were a rare finding.In B cells the penetration of collagen fibrils through the cell membrane was frequently observed (Figs 5, 6).As can be seen in Fig. 6, these fibrils arose from the content of transport vacuoles travelling towards the cell membrane.Tropocollagen was spilled onto the surface and the following polymerization produced collagen fibrils which showed the signs of periodicity (Figs 5,6). | 12 |
34721866 | Synovial matrix This is generally characterized as an intercellular substance, originating from mesenchyme, which provides a specialized environment for synovialocytes.As with other mesenchymal tissues, it consists of the ground fibrillar substance and the ground amorphous substance. In the synovial membrane of the adult pig, the ground fibrillar substance was made up of two types of fibrils.First, typical collagen fibrils, 60-100 nm thick and several I'm long, which were branched and showed a periodicity of 64 nm.Second, aperiodic fibrils about 50 nm in thickness and 0.1 I'm in length or, on rare occasions, longer.These two kinds of fibrils were observed in the cisternae of the granular endoplasmic reticulum of both types of synovialocytes.Similar findings have been made by Wassilev (1975), Ghadially (1983) and Horky (1984 in the synovial membranes of various other mammalian species. The ground amorphous substance was composed of the protein-hyaluronic acid complex and sulphonated mucopolysaccharides.The two components were visualized by electron microscopy as finely granulated, medium-osmiophilic matter present among collagen fibres.They both probably migrated into the synovial fluid, taking part in lubrication processes (Hills and Butler 1984). In the adult pig, the synovial matrix varied in arrangement from place to place in relation to the distribution of synovialocytes.In areas where the cells were situated near the surface of the synovial membrane (Fig. 1) the surface consisted largely of parallel bundles of aperiodic filaments.These were embedded in a relatively small amount of ground amorphous substance which produced a layer at the boundary of the articular cavity.More fibrillar structures, i. e. irregularly running aperiodic filaments and collagen protofibrils, thin collagen fibrils showing periodicity, were seen in the amorphous substance at the side of synovialocytes or in deeper layers of the synovial membrane.Aperiodic filaments in close contact with the synovialocyte cell membranes were frequently observed (Figs 1,2,3). In the regions where the cells oceurred in deeper layers and the surface of the synovial membrane was formed mostly by their cytoplasmic projections, the surface showed a different arrangement.Numerous cytoplasmic projections were present in the ground amorphous substance while the fibrillar component was considerably reduced in amount (Fig. 2).Aperiodic filaments were present in low numbers and collagen fibrils near the surface were a rare finding.Some of the cytoplasmic projections were unmasked and got in direct contact with the articular cavity (Fig. 2).A large number of collagen fibrils could be found close to the cell membranes of synovialocytes facing the deeper layers of the synovial membrane (Figs 2, 4).In the two regions of synovial membrane, collagen fibrils were observed behind the basement membrane. | 13 |
34721866 | Discussion The synovial membrane is a tissue originating from mesenchyme.It provides lining for the articular fissure.Its structure is generally that of connective tissues but the cells, which form a monolayer or a mUltilayer cover, differ from the other cells of connective tissue in morphology and function.The synovial membrane arises from the original skeletal blastema during development (Andersen 1964; Stoff and Effendy 1985).The cells covering the synovial membrane surface' ate regarded as modified mesothelial cells even though they do not produce a continuous layer and are not interconnected by intercellular bridges such as desmo-• somes or zonulae occludentes, etc.It has been known that synovial cells are, segregated.from the surface and are replaced with cells proliferating from the' subsynovial layer; this is particularly obvious during inflammatory diseases.(Ghadially 1983).As shown by experiments using 3H-thymidine, the labelled cells appear in the normal synovial membrane only occasionally.Their numbers,.however, increase following experimental inflammation or partial synovectomy, . .which was demonstrated by Schulitz (1974) in the synovial membrane of rabbits.A typical feature of this tissue is the absence of neural endings.Occasionally' seen nerve fibres are autonomous fibres of vascular adventitia (Halata and Groth 1976; Knight and Lewick 1983). The function of synovial membrane has been studied thoroughly.Evidenceobtained up to the present suggests two principal roles: (1) production of synovial fluid (2) exchange and removal of synovial fluid and cell detritus related to the arti-• cular cavity. The articular fluid is a specific plasma transudate enriched with substances'.excreted by type B synovialocytes; this was demonstrated by Swann in 1978• and recently by other authors who studied relationships between the synovial fluid and the articular cartilage with respect to nutrition and, particularly, to• the mechanics of articular movement (Sokoloff 1980; Swann et al. 1981, 1984,. 198; Hills and Butler 1984; Sabiston and Adams 1989) or in association. with the role of enzymes in degradation processes (Krane and Amento 1983;Markowitz 1983;Dingle 1984;Gangel 1984).The finding that hyaluronicacid (non-sulphonated glycosamine glycan) is transferred to the synovial fluid was made a long time ago (Lever and Ford 1958) and re-confirmed recently (Hilbert et al. 1984).All the components of synovial membrane, excluding~ digestive enzymes, are involved in the nourishment of the articular cartilage and in lubrication qualities of the synovial fluid.The other role of the synovial membrane, i. e. removal of cell detritus, has been reported by Ball et al. (1964) and similar data were presented by Horky et al. (1974) in hemarthrosis. The synovial membrane surface has a villous character.In different areas• ist appearance varies in relation to the state of the membrane under either physiologic or pathologic conditions.This was studied by scanning electron microscopy by Kondoh (1973), Gaucher et al. (1976) Horky (1981) and Ghadially (1983). Synovial cells are embedded in the matrix synoviaIis whose density ranges• from low to medium values.The intercellular matter includes typical collage fibrils showing periodicity, aperiodic collagen fibrils and fine aperiodic filaments situated in the amorphous substance (Meachim and Stockwell 1979; HorkY-1981HorkY- , 1984, 1989ab), 1989ab).Collagen fibrils of typical appearance are present in greater -amounts in the deeper parts of the membrane, while near the surface the predo-_ minant fibrillar components are collagen fibrils without periodicity and periodic filaments.One of the explanations has suggested that collagen fibrils in deep Jayers are gradually disintegrated down to aperiodic filaments (Ghadially 1983) but from the data on functions of type A synovialocytes and from the morpho--logical evidence on the ability of fibrills to pass through the cell membrane (H 0 r-.kY 1981, 1984, 1~89ab) it seems more probable that this process takes the other way round. Although the microscopic and submicroscopic structures of the synovial membrane have been studied in a great range of mammalian species (Langer and Huht 1960;Barland et al. 1962;Cutlip and Cheville 1973;Fell et al. 1976;Linck and Porte 1978;Okada et al. 1981;Horky 1981Horky , 1984)), the _ information on the synovial membrane in the pig is scarce.The in vivo structure was described by Fell et al. (1976) who were also interested in the behaviour •of synovialocytes in vitro.The ultrastructure of the porcine synovial membrane has only been studied during the prenatal and early postnatal periods (Horky 1989ab).Thus the observations presented here can be compared with our pre-• vious results obtained in the pig or with those concerning the bovine synovial membrane. A characteristic feature of the porcine synovial membrane under study was • a mixed population of fully differentiated type A and type B cells and transient A -B cells.The distribution of the respective types was similar to that reported in the adults of other mammalian species excluding the rat (Wassilev 1975).To distinguish between them is not difficult and it is based on distinct appearances of the cytoplasm related to the presence of organelles and secretory granules.These bodies have been described in adult animals of several mammalian species .(Ghadially and Roy 1969;Linck and Porte 1978;Graabaek 1984) and in the prenatal and early postnatal periods of different mammals including man - (Horky 1981(Horky , 1984, 1989ab), 1989ab).They have been given various names and the opinions on their origin, composition and function are diverse.The results published by Linck and Porte (1978) and particularly Okada et al. (1981) and Graabaek ,(1984and Graabaek ,( , 1985) ) demonstrated that in formation of secretory granules a major role is played by the granular endoplasmic reticulum and Golgi complex.Okada et al. (1981) and also Graabaek (1984Graabaek ( , 1985) ) showed that, in contrast to A-cell .lysosomes, the secretory granules of B cells do not contain acid phosphatase but have mucopolysaccharides and glycoproteins bound to a protein carrier, which could be proved by protein digestion. Apart from the distinct types of synovialocytes, our observations included also cells where the presence of cellular organelles was indicative of A cells but • which also contained secretory granules.These fi ldings supported our earlier view (Horky et al. 1974(Horky et al. , 1975(Horky et al. , 1981(Horky et al. , 1984, 1989ab) , 1989ab) as well as the views of other authors (Fell et al. 1976;Linck and Porte 1978;Okada et al. 1981) that trans--formation of type A and B cells and vice versa is possible and depends on the :species of the animal and its age and the physiologic state of the joint (Cutlip and Cheville 1973).In addition, Linck and Porte (1978) and Fell et al . • (1976) ascertained that this reflects functional flexibility of the synovial membrane ,and not the loss of cell function. In comparison with the porcine foetal synovial membrane, the synovialocytes ;under study regularly involve intracytoplasmic filaments (Lazarides 1980; Horky 1984.In adulthood, type A cells of synovial membrane form basement membranes which are absent in the A cells of the prenatal and perinatal periods and in B cells (Horky 1989ab).As seen previously in cattle in the prenatal and perinatal periods (Horky 1989) and in the same periods in the pig (Horky 1989ab), collagen fibrils were able to pass through the cell membranes of B cells.Thus the designation of B cells as S (secretory) cells, based on production of various substances, is fully justified. | 14 |
216464946 | ERROR: type should be string, got "https://doi.org/10.1590/1981-6723.11519 Abstract The aim of this research was to evaluate the influence of sweet potato peel flour (SPPF) on the physicochemical, technological and sensorial characteristics of bovine hamburger. Four hamburger formulations were prepared added SPPF: F1 (0%), F2 (0.75%), F3 (1.5%) and F4 (2.25%). The flour was characterized by high levels of minerals, carbohydrate and dietary fiber, which improved the nutritional profile of the hamburger. There was an increase in moisture retention and shrinkage, as well as a reduction in fat retention and cooking yield, as the level of SPPF addition increased. The addition of flour in the product significantly reduced ( p < 0.05) the values of L* , a* and b* . Similar acceptability to the standard sample was checked for the hamburger with the addition of up to 1.5% SPPF. However, all formulations had an acceptability index greater than 70%. It is concluded that SPPF is a potential ingredient to be added in bovine hamburger, improving nutritional and technological parameters and with low influence on the sensorial gordura e no rendimento da cocção, conforme se elevou o nível de adição de FCBD. O acréscimo de farinha no produto reduziu significativamente ( p < 0,05) os valores de L* , a* e b*. Aceitabilidade similar à amostra padrão foi verificada para o hambúrguer com adição de até 1,5% de FCBD. Contudo, todas as formulações apresentaram um índice de aceitabilidade superior a 70%. Conclui-se que a FCBD é um ingrediente com potencial para adição em hambúrguer bovino, melhorando parâmetros nutricionais e tecnológicos e com baixa influência nas características sensoriais." | 1 |
216464946 | Introduction Meat and meat derivatives are much-appreciated types of food by consumers as part of the regular diet. In addition, they present a positive nutritional profile, mainly regarding protein content and quality (Utrera et al., 2014). However, high meat consumption, especially red and processed meat, has been linked to the increased risk of developing diseases such as cancer (Qu et al., 2013;Zhu et al., 2013), stroke (Chen et al., 2013), diabetes mellitus type 2 (Feskens et al., 2013) and cardiovascular diseases (Abete et al., 2014). Some meat products are notable for their high consumption, among them are hamburgers, meatballs and sausages. This fact is mainly due to the practicality, ease of preparation, besides being very tasty and generally are financially affordable to the population. Currently, the seek for safer, healthier and tastier products is eminent in the world's population. In this case, there is an encouragement to the development of products that offer a better nutritional profile (De Smet & Vossen, 2016) and which promotes a better quality of life and a reduction in the risk of development of pathologies (Domingo & Nadal, 2016). The application of alternative ingredients, such as the peel of vegetables, can be considered a potential strategy since it can increase the value-added to the product. Annually, 95% of by-products from vegetables (peels, stems, seeds and leaves) are discarded during preparation and processing (Melikoglu et al., 2013). Besides the waste of food, this fact contributes to the increase of organic waste, which damages the environment. Research has shown that the nutritional content of vegetable peels is very beneficial for human consumption. It may also contain more nutrients than the pulp itself, such as vitamins, minerals and fibers (Moo-Huchin et al., 2014). Sweet potato is a tuberous root belonging to the family Convolvulaceae. It is widely cultivated in several countries (Shan et al., 2013), and China is the largest producer of sweet potatoes in the world (Food and Agricultural Organization of the United Nations, 2014). In 2012, the annual world production of sweet potatoes was approximately 108,004 million tons, concentrated in regions such as Asia (71.6%) and Africa (16.9%) (Food and Agricultural Organization of the United Nations, 2014). In Brazil, sweet potato ranks third place among the most consumed tuberous roots and seventeen in total crop production (Food and Agriculture Organization, 2015). The pulp is mainly composed of carbohydrates, protein, minerals and fibers (Grace et al., 2014). The peel contains even higher levels of fiber, vitamins and minerals such as potassium, magnesium and folate (Food and Agriculture Organization, 2015). The cultivars of sweet potato peel and purple pulp may contain considerable levels of acylated anthocyanins and other phenolics, which have antioxidant and anti-inflammatory functions (Grace et al., 2014). Despite this, consumption of sweet potato peel is not frequent. In that respect, the addition of this by-product to meat products, such as hamburgers, could help increase nutritional value and reduce organic waste in the environment. In addition to the nutritional question, research has already shown that the addition of sweet potato peel in meat products can maintain or even improve technological aspects such as texture and flavor (Tokusoglu & Swanson, 2014;Mehta et al., 2015). In this context, the objective of the present research was to evaluate the influence of the addition of sweet potato peel flour (SPPF) on the physicochemical, technological and sensorial characteristics of bovine hamburger. | 2 |
216464946 | Sweet Potato Peel Flour (SPPF) elaboration Purple sweet potatoes (70 kg) were used, showing a good visual appearance, smooth surface without imperfections and medium size. The whole sweet potatoes (Ipomoea batatas L. (Lam.)) were washed and immersed in a sodium hypochlorite solution, with a proportion of 8 ml for each liter of water. After 15 minutes, the tubers were rinsed again under running water. The peels (2 mm thick) were manually removed with the aid of a knife and dried in a dehydrator with forced air circulation (Pardal ® , PE 60, Brazil) at 60 °C for 24 hours. Peels were grounded in a mill (Tecnal ® , Tec mill TE-633, Brazil), yielding 1.4 kg of flour. The product was packed and stored at -18 °C until analyzes were carried out. | 3 |
216464946 | Beef patties processing and cooking Four formulations of hamburger were prepared, containing three independent replicates of each treatment: beef (shoulder clod) (F1: 77.9%, F2: 77.1%, F3: 76.4% and F4: 75.7%), SPPF (F1: 0%, F2: 0.75%, F3: 1.5% and F4: 2.25%), ice flakes (15%), pork fat (5%), sodium chloride (1.5%), onion powder (0.2%), garlic powder (0.2%) and black pepper. The percentages of each ingredient were defined by means of preliminary sensorial tests carried out with the product. To elaborate the hamburgers, the meat (approximately 10 kg) was ground in a meat grinder (C.A.F. ® , Brazil), on a 3 mm disk and with a temperature around 4 °C. Subsequently, the ground beef was then homogenized in a commercial blender (Super Cutter Sire ® , Brazil) for 1 minute at 9 ± 1 °C. The onion, garlic, pepper, sodium chloride, ice flake and pork fat were added to the mixture and homogenized again for 3 minutes at 9 ± 1 °C. SPPF was incorporated into the dough and homogenized for an additional 3 minutes at 9 ± 1 °C. Additional levels of ground beef and SPPF differed in each formulation as described above. The resulting dough of each formulation was shaped into hamburgers (100 g, 10 cm in diameter and 1 cm thick) using a hand-fed hamburger (Picelli ® , HP 128, Brazil). The products were stored in plastic bags of low-density polyethylene and frozen in a conventional freezer (-18 °C) for 10 days. The frozen hamburgers were grilled on an electric plate with grill on the upper and lower sides (Britania grill ® mega 2N, Brazil) heated to 200 °C. The internal temperature of the hamburger was controlled by a digital thermometer (Tp 101 ® , Brazil) until reaching 71 °C at its geometric center (American Meat Science Association, 2015). The average cooking time was 8 to 10 minutes. | 4 |
216464946 | Physicochemical composition All analyses were performed on three replicates in triplicate for SPPF and for cooked hamburgers. Moisture, ash, protein, fat and dietary fiber content were determined by the Association of Official Analytical Chemists (2011). The moisture content was determined by drying in a greenhouse (105 ± 2 °C). Fat content was determined according to the Soxhlet method, using petroleum ether. Protein was analyzed according to the Kjeldahl method. Factor 6.25 was used for the conversion of nitrogen to crude protein in hamburger and SPPF respectively. Ash was performed by a muffle furnace. Total, soluble and insoluble dietary fiber was determined by the enzymatic method. The carbohydrate content was evaluated by means of theoretical calculation (by difference) in the results of the triplicates, according to the Formula 1: The total caloric value (kcal) was calculated theoretically using Atwater factors (Atwater & Woods, 1896) for lipid (9 kcal g -1 ), protein (4 kcal g -1 ) and carbohydrate (4 kcal g -1 ). Water activity (Aw) hamburgers were used per treatment, evaluated in five different points of the hamburger. The color was evaluated by the system of the Commission Internationale de L'Eclairage (CIE), lightness (L*), redness (a*), yellowness (b*), colorimeter reading (Konica Minolta ® , Chroma Meter CR 4400 model, Japan) with illuminating calibration D65 and angle of observation 10º, previously calibrated. | 5 |
216464946 | Technological analyses Five hamburgers from each formulation were cooked in the same procedure as mentioned previously then cooled to room temperature at 23 °C for 2 h. The following cooking characteristics were evaluated: cooking yield (2) and fat retention (3) (Murphy et al., 1975), shrinkage (4) (Berry, 1992) and moisture retention (5) (El-Magoli et al., 1996). All experiments were done in triplicate. The hamburgers were measured according to the following Equations 2-5: | 6 |
216464946 | Sensorial analyses Participated in sensory analyses 65 untrained volunteer subjects, hamburger usual consumers. Consumers had aged between 18 and 50 years and were recruited among students and staff of Universidade Estadual do Centro-Oeste, Guarapuava, Paraná, Brazil. For conducting the sensory test, hamburgers have been cooked as previously described. All samples were evaluated by means of an acceptance test using a nine-point hedonic scale, with extremes ranging from dislike extremely (1) to like extremely (9) (Meilgaard et al., 1999). Attributes related to appearance, aroma, flavor, color and texture, beyond overall acceptance were evaluated. For the purchase intent test a 5-point attitude structured scale was used, varying from definitely would not buy it (1) to definitely would buy it (5) (Meilgaard et al., 1999). The sensory Acceptability Index (AI) was calculated by multiplying the average score reported by consumers to the product by 100, dividing the result by the maximum average score given to the product within the hedonic scale of 9.0 points. Each sample was served to consumers in white plates coded with randomly selected 3-digit numbers in monadic form and using balanced design (Macfie et al., 1989). Sensory evaluations were performed by consumers under fluorescence lighting. After consuming each sample, the consumer was instructed to drink water for palate cleansing. Samples were evaluated in triplicate in a separate session. | 7 |
216464946 | Ethical issues The study was approved by the Ethics in Research Committee of UNICENTRO, Brazil, under the case number of 608.950/2014. Sweet potato peel contains high carbohydrate and fiber content when compared to beef which is exempt in its composition (United States Department of Agriculture, 2014). Studies have shown that adequate fiber consumption reduces the risk of developing pathologies such as cardiovascular disorders (Mirmiran et al., 2016), systemic arterial hypertension (Evans et al., 2015), diabetes mellitus (Wu et al., 2015), among others. Moisture, ash, carbohydrate and fiber contents increased with the addition of SPPF. The highest moisture content of the SPPF hamburger is explained by the water retention property of the fibers (Célino et al., 2014), as previously reported. In addition, fibers interact with proteins of the meat, resulting in a network that prevents the translocation of water from the product to the surface (Song et al., 2016). The higher ash, carbohydrate and fiber content in F2, F3 and F4 are due to the higher amount of these nutrients present in SPPF compared to meat. Similar results were verified after the addition of orange peel flour (5%) in bovine hamburger (Mahmoud et al., 2017). Protein, lipid and caloric contents were lower for SPPF-added hamburgers since SPPF contains lower levels of these nutrients compared to meat. These results corroborate with other studies evaluating the addition of poppy seed (Gök et al., 2011) and orange peel flour in bovine hamburger (Mahmoud et al., 2017) reduces protein and lipid content. There was no significant difference (p > 0.05) between the pH and Aw results of the formulations, as reported in the literature (Longato et al., 2017). The instrumental color results of cooked hamburgers are presented in Table 2. less red and yellow, since the sweet potato peel has a light brown color. In addition, the sweet potato peel has catalytic chelating metals, such as iron and zinc, which favor oxidation of lipids and proteins present in meat (Ahn et al., 2002;Lund et al., 2007;Andrés et al., 2017). These compounds alter the color of the product, which reduces consumer acceptability (Jha et al., 2007). Garcia et al. (2009), who evaluated hamburger with dry tomato peel (1.5% to 6.0%), reported similar results. | 9 |
216464946 | Technological analyses The results of the cooking characteristics of hamburgers are shown in Table 3. The shrinkage and moisture retention increased after the increase of SPPF in the hamburger, due to the high fiber content of the sweet potato peel, which retains water in the product, increasing the succulence (Anderson & Berry, 2001). However, there was a reduction in cooking yield (p < 0.05) and fat retention of SPPF hamburgers, corroborating with the literature (Gök et al., 2011). The preferential bonding of the fibers by water in detriment of fat may explain these findings (Anderson & Berry, 2001), because the fibers form gels in aqueous solution, a process called myofibrillar protein gelation (Cordeiro, 2011). | 10 |
216464946 | Sensorial analyses The results of the hamburger sensory test added at different levels of SPPF are described in Table 4. Table 4. Sensory scores (mean ± standard error) obtained for the hamburger with the addition of different levels sweet potato peel flour (SPPF). (2.25%) reduces product acceptance, due to the residual and bitter taste of phenolic compounds present in large quantities in sweet potato peel (Anastácio et al., 2016). Moreover, the addition of SPPF in the hamburger modified the texture of the dough making it more brittle, due to sweet potato peel high fiber content. Fiber hygroscopic capacity may explain this effect since they retain water inside the product (Célino et al., 2014). All formulations showed high acceptance rates (≥ 70%), which demonstrate good sensorial acceptance of the products (Corradini et al., 2014). Thus, it is demonstrated the feasibility of using SPPF as an ingredient in hamburger, which favors the consumption of healthier foods by the population. | 11 |
44236269 | The electronic structures of nine dihalobenzenes (C6H4FX; X = Cl, Br, I) have been studied by UV photoelectron spectroscopy (UPS) and assigned by comparison with the reported spectra of monohalobenzenes (C6H5X; X = Cl, Br, I). and quantum chemical calculations. Our results show that the fluorine substituent modifies energies of πand halogen lone pair orbitals to a significant degree depending on its location (topology). We also demonstrate that the inductive effect of fluorine atom on the benzene ring can be readily observed and interpreted. | 1 |
44236269 | INTRODUCTION In their recent work Antal et al. considered the influence of through-bond and through-space interactions on the shape of molecular electron density. 1 The authors used theoretical method of shape analysis to provide quantitative measure of through-bond (TB) and through-space (TS) effects.Because of our long standing interest in the analysis and systematization of halogen substituent interactions in halobenzenes, we provide in this note some experimental results which can be useful for assessing the validity of theoretical concepts attached to TB and TS effects.We used accurate vertical ionization energies (which are good approximations of orbital energies) obtained from UV photoelectron spectroscopy (UPS) as the quantitative probe of TB and TS effects.Antal et al. 1 selected substituted styrene derivatives as their test case.However, we use smaller dihalobenzenes, because they exhibit well resolved spectral bands thus allowing accurate ionization energies to be measured; this would not have been possible with larger styrene derivatives where the bands can be expected to overlap in the spectra.Furthermore, the unambiguous spectral assignments for our molecules can be readily obtained using the empirical arguments.These arguments can thus support any conclusions drawn from theoretical calculations. | 2 |
44236269 | EXPERIMENTAL Samples of the compounds studied in this work were obtained from Sigma-Aldrich.The photoelectron spectra (Figures 1-9) were recorded on the Vacuum Generators UV-G3 spectrometer and calibrated with small amounts of Xe gas which was added to the sample flow.The spectral resolution in HeI spectra was 25 meV when measured as FWHM of the 3p -1 2 P 3/2 Ar + ← Ar ( 1 S 0 ) line.The vertical ionization energy values have been determined at the band maxima.All the samples were liquids and their spectra were recorded at room temperature.The measured spectra were reproducible and showed no signs of decomposition e.g.no sharp peaks corresponding to possible small molecules (decomposition products) were observed.The spectra were reproducible over long time intervals.The quantum chemical calculations were performed with Gaussian 09 program 2 and included full geometry optimization of neutral molecules using B3LYP functional, 6-31G* basis set on all atoms except iodine where Stuttgart effective core potentials was used. 3he vibrational analysis confirmed that the resulting geometry was the true minimum (no imaginary frequencies).Subsequently, the optimized DFT geometry was used as the input into the single point calculation using the outer-valence Green's function (OVGF) method and the same basis sets. 3This method obviates the need for using Koopmans approximation and provides vertical ionization energies with typical deviation of 0.3-0.5 eV (depending on the size of the basis set) from the experimental values. | 3 |
44236269 | RESULTS AND DISCUSSION The photoelectron spectra are shown in Figures 1-9 and their assignments are summarized in Table 1.We shall briefly discuss the generic assignment of the spectra first and then focus on the substituent effects which can be discerned from the spectra. It is well established that in the UPS spectra of monohalobenzenes the lowest ionization energies correspond to π-ionizations (π 3 and π 2 orbitals) from HOMO and SHOMO orbitals.The next two bands at higher ionization energies correspond to halogen lone pairs of Cl, Br or I.][6][7] This well established, empirical assignment (Scheme 1) allows us to readily assign our spectra as well (Table 1) and furthermore, it is also supported by the results of OVGF calculations (Table 1). Before discussing inductive and resonance effects we recall several of their basic characteristics.The inductive effects operate via the network of σ-bonds and are therefore of predominantly TB type.The inductive effects are electrostatic in nature.The resonance effects depend on orbital overlap (often involving π-orbitals).Therefore the resonance effects involve predominantly (but not exclusively) TS interactions.The inspection of Table 2 shows several distinct trends regarding the influence of substituent topology on the electronic structure of dihalobenzenes.We also recall that the fluorine substituent acts on the aromatic system mostly in an inductive manner.We discuss next the orbital energy effects deduced from the spectra. | 4 |
44236269 | 1. Energy splitting between π-orbital energies Δπ (Δπ = π 3 -π 2 ) can be taken as an indicator of the π-electronic structure of the molecule.Δπ increases in para-substituted C 6 H 4 FX and decreases in meta-and ortho-isomers (all measured relative to the Δπ in the corresponding C 6 H 5 X; X = Cl, Br, I).The energy splitting between two halogen lone pairs in the C 6 H 4 FX molecules (Δn X ) remains virtually unchanged in the ortho derivatives, but is reduced in the meta-and para-isomers as shown in Table 2 (changes are measured again vs. Δn X in the corresponding C 6 H 5 X; X = Cl, Br, I). | 5 |
44236269 | 2. In order to monitor the inductive effects we calculated average values of π and n X orbital ionization energies which we designate as <π> and <n X >, respectively.The results in Table 2 show the effect of fluorine on <π>; in C 6 H 4 FX <π> does not depend markedly on the position of the fluorine substituent (the variations are < 0.1 eV).The C 6 H 4 FI isomers are an exception.In these molecules the location of fluorine substituent has large influence on the <π> values; the strongest effect occurs in the ortho-isomer.Similar, limited influence of topology of fluorine substitution is evident for halogen lone pairs <n X >.However, the C 6 H 4 FI isomers once again show stronger dependence of <n X > on the position of the fluorine atom. The qualitative rationalization of these trends can be made with reference to the Scheme 1 as follows. Ad 1.In para-C 6 H 4 FX isomers the electron rich fluorine atom is located close to the maximum of the electron density of the HOMO, but not of SHOMO π-orbital.This can be expected to destabilize the HOMO more than the SHOMO and thus increase the Δπ value.In the ortho-and meta-C 6 H 4 FX isomers the electron rich fluorine atom is situated close to the maximum of electron density of the SHOMO orbital; this orbital is therefore destabilized in preference to the HOMO which results in the reduced Δπ splitting.When considering the effect of topology of fluorine substituent on the X lone pairs in C 6 H 4 FX we need to recall that the fluorine substituent acts inductively via σ-bond framework (TB interaction) and mostly affects the in-plane (n Xσ ; σ-symmetry) halogen lone pair orbital rather than the out-of-plane (n Xπ ; π-symmetry) lone pair.In general, the C-F bond dipole will tend to pull the halogen lone pair density away from X (in the direction of dipole's spatial orientation) thus stabilizing the corresponding n Xσ and reducing Δn X .In ortho-C 6 H 4 FX isomers we noticed two opposing effects.The C-F bond dipole still pulls electron density away from n Xσ , stabilizing it.However, due to the spatial proximity between X halogen lone pair and fluorine lone pair in ortho-isomers we also have TS interaction between two fully occupied orbitals which leads to the destabilization of the lone pair with higher energy which is n Xσ .The net result is that there is no significant change in n Xσ orbital energy and hence no change in Δn X .On the other hand, in meta-and para-C 6 H 4 FX isomers, there is no corresponding TS interaction and C-F bond dipoles tend to pull the electron densities away from the in-plane X lone pairs thus stabilizing the n Xσ orbital and reducing Δn X . Ad 2. The reason why the <π> and <n X > values in C 6 H 4 FI are influenced by the positions of fluorine substituents more than in other dihalobenzene molecules is due to the large polarizability (and the low electronegativity) of iodine atom compared to the chlorine or bromine atoms. | 6 |
56286785 | Egyptian Academic Journal of Biological Sciences is the official English language journal of the Egyptian Society for Biological Sciences, Department of Entomology, Faculty of Sciences Ain Shams University. Histology& Histochemistry Journal include various morphological, anatomical, histological, histochemical, toxicological, physiological changes associated with individuals, and populations. In addition, the journal promotes research on biochemical and molecularbiological or environmental, toxicological and occupational aspects of pathology are requested as well as developmental and histological studies on light and electron microscopical level, or case reports. www.eajbs.eg.net Provided for non-commercial research and education use. Not for reproduction, distribution or commercial use. | 1 |
56286785 | Experimental animals: Male adult albino mice (Mus musculus) obtained from National Research Centre, Cairo, was used in all experiments conducted in this study.At the beginning of each experiment the mice age between (7 to 10 weeks) and weighing between (20-30 gm) were kept in cages under standard conditions, i.e. a well ventilated room and a controlled regimen of fluorescent light (light for 12 hours and dark for 12 hours) at the Animal House of Zoology department, Faculty of Science, Suez Canal University.Mice were housed in plastic cages, wire topped with sawdust bedding.The sawdust bedding of the mouse boxes was changed weekly and the cages were cleaned and sterilized.Mice were fed on standard diet and tap water was given ad libitum.They were acclimatized to their place for one week before the experiment.Animals were randomly allocated into four separate experiments.The animals were used according to the guidelines of the Committee on Care and Use of Experimental Animal Resources (Suez Canal University, Egypt) and all efforts were made to reduce the number of animals used and their suffering. | 3 |
56286785 | Experimental design: Animals were divided into 4 groups (a control group and three treated ones) of 5 male mice each.Three groups were injected intraperitoneally with 5-FU (5, 10 and 15 mg/kg) for five consecutive days at intervals of 24 hr.Mice of the control group were injected with injectable water.All animals were sacrificed on day 35 following the last injection.Post treatment sampling at 35 days was chosen for the study to allow the germ cells, which were exposed at late spermatogonial stage to the drug (5-FU) to reach the caudal epididymis. | 4 |
56286785 | Histological preparation: Animals were anesthetized using chloroform, dissected and tissues samples of testes were taken out.Tissue samples were fixed in Bouins fluid for 48 hr.The fixative was changed twice during 48 hr.The samples were then washed several times in 70% ethyl alcohol to remove the excess fixative, dehydrated using ascending series of alcohols (80%, 90%, 100% and 100%), cleared in Terpineol and embedded in paraffin wax.Tissue blocks were sectioned at 5 µ thickness and stained with Harris Haematoxylin and Eosin (Mallory, 1944). | 5 |
56286785 | Sperm sampling and staining: The epididymides were excised and minced in 1 ml of 0.9% physiological saline.The contents were gently pipetted or squeezed five to six times up and down in a 5 ml pipette.The sperm solution was filtered through a nylon cloth to remove tissue fragments.A small drop of the cell suspension was put on the end of a clean slide and spread by pulling the material behind a clean glass cover held at an angle of 45 degrees.The slides were air dried without fixation for about 24 hr.Slides were stained with 1% Eosin-Y (aqueous) for 30 minutes followed by two rinses in distilled water.Slides were then left to air dry and cleared in two changes of Xylene, 5 minutes each.Five slides were prepared for each mouse.Sperm smears were examined by light microscopy.For each mouse, 800 sperms were examined and morphological abnormalities of sperm head and tail were recorded according to the criteria of EL-Nahas et al. (1989).Abnormal sperm morphology is classified as defects in the head, midpieces and tail (Burruel et al., 1996). | 6 |
56286785 | RESULT Testes of control mice were histological normal (Fig. 1).Testes of treated mice showed histological altered seminiferous tubules, at all doses.This alteration was in form of sloughing (loss) of immature germ cells into the tubular lumen.Some of the tubules showed haloapperance in the round spermatids.Spermatids of some tubules were observed forming round multinucleated giant cells (Fig. 2). 5-FU treatment affected the percentages of morphologically abnormal sperms at all of the three tested doses.Various morphological sperm abnormalities were observed in control and treated animals.Morphological abnormalities of mice sperms induced by 5-FU treatment, in the present study, were grossly headed sperms, quasi-normal headed sperms, angular midpiece sperms and bended tailed sperms (Fig. 3A-C). Figure ( 4) showed graphically the mean values of morphologically abnormal sperms for the four studied groups while Figure ( 5) showed the linear trendline for the four studied groups which revealed that the mean values of the morphologically abnormal sperms increased as the drug doses increased. The ANOVA test showed that the statistical differences between the control group and the 5-FU treated groups (5, 10 and 15 mg/kg) were statistically highly significant (p= 0.000267 at the level of significance of 0.05).The multiple comparisons between the four studied groups using Post Hoc Tukey test revealed that the statistical differences between the control and each of the studied drug doses (5, 10 and 15 mg/kg) were highly significant (p = 0.004, 0.003 and 0.001 respectively at level of significance of 0.05).However, the statistical differences among the drug doses 5, 10 and 15 mg/kg were insignificant. | 8 |
56286785 | DISCUSSION At all used doses, the testes of the treated mice showed some histologically altered seminiferous tubules, whereas other seminiferous tubules were not affected.The fact that some of the seminiferous tubules were altered and other tubules were not, had been supported by Meistrich et al. (1982) who mentioned that the intracellular half-life of the drug is 7 to 9 days so that the stem cells may be triggered into cycle by administration of 5-FU while the drug is still active.Histologically altered seminiferous tubules showed sloughing of the germinal epithelium.This is similar to studies in rats by D 'Souza and Narayana (2001) who found sloughing of the germinal epithelium in the lumen after injecting 10, 50 and 100 mg/kg 5-FU intraperitoneally. In the present study, multinucleated giant cells and haloapearence spermatids observed in seminiferous tubulus of the treated mice are in agreement with the findings of Narayana et al. (2000) who found multinucleated giant cells in the seminiferous tubulus lumen after injecting 100 mg/kg 5-FU intraperitoneally.Regarding the results of sperm morphology assay, this work revealed a statistically significant increase in the percentage of morphologically abnormal sperms at all used doses.Such percentages were 39.5%, 41% and 47.2% for 5, 10 and 15 mg/kg 5-FU respectively, as compared with 15.1% for the control animals.The differences between the control group and the treated groups were statistically significant (p<0.05),but the differences in the abnormal sperm count between the treated groups turned to be statistically insignificant.This is not in agreement with the findings of Choudhury et al. (2002) who used a single intraperitoneal injection of 5, 10 and 15 mg/kg 5-FU and carry out sperm morphology assay at week 8 post-treatment.He reported high percentages of abnormal sperm, but were not statistically significant.He attributed this to the gradual decline in the transmission of the induced cytogenetic toxic effects of 5-FU from spermatogonia to sperm, due to gradual elimination of the grossly affected spermatogonial cells during the course of spermatogenesis. Morphological abnormalities of mice sperms induced by 5-FU treatment, in the present study, were grossly headed sperms, quasi-normal headed sperms, angular midpiece sperms and bended tailed sperms. According to Wyrobeck (1984) the significant increase in the number of morphologically abnormal sperm has been associated with infertility.In the three used doses (5, 10 and 15 mg/kg) of 5-FU quasi-normal headed sperms were observed in the examined sperm smears.Quasi-normal head defects do not seem to affect the motility of spermatozoa but significantly reduce the in vitro and the in vivo fertilizing capacity (Jeyendran et al., 1986). The quasi-normal head may be due to the action of 5-FU on the genes responsible for expression of acrosomes characteristics (Topham, 1980).Sperms with abnormal tail either with coiled tail or bended tail were observed.These abnormalities affect the motility of the sperm.Menkeld et al. (1990) related tail coiling to sperm aging.Sperms with bended midpiece were recorded.Menkeld et al. (1990) suggested that bended midpiece had grown from wrong centriole.Sperm cell morphology is genetically controlled by numerous autosomal and sex linked genes (Krazanowska, 1976).Hence, formation of abnormal sperm population in the present study is very likely due to the mutagenic effects of 5-FU on the specific gene loci of germ cell chromosomes involved in the maintenance of normal sperm structure.These mutagenic effects of 5-FU are primarilary caused by the direct cytotoxicity of germ cells during spermatogenesis.The cytotoxicity is caused by the antimetabolic activity of 5-FU through the inhibition of thymidylate synthestase and the erroneous incorporation into RNA and DNA (O`Dwyer et al., 1987 andPinedo andPeters, 1988). We conclude that even small dose of 5-FU affect the patient fertility by disturbing the testes histology and sperm morphology. | 9 |
56286785 | Fig. 2 : Fig. 2: Testes section of mice injected intrapertoneally with 15 mg/kg 5-FU for 5 consecutive days and left for 35 days showing sloughing (S) of immature germ cells into the tubular lumen.Some of the tubules showed haloapperance in the round spermatids (h).Spermatids of some tubules were observed forming round multinucleated giant cells (GC) (HE, X 200). | 11 |
35371240 | A simple, precise, accurate, high reproducible and economical visible spectrophotometric method of analysis for the synthesized rasagiline hemitartrate was developed and validated. The proposed method involves diazotization of sulphanilic acid under acidic conditions in presence of sodium nitrite, followed by its coupling with rasagiline in alkaline medium. The absorption spectra of the yellow colored chromophore formed between rasagiline and positive diazonium ion has absorption maximum at 440 nm. The linear regression analysis data for the calibration plot showed good linear relationship (r = 0.99937) with in the concentration range of 0 – 10 μg mL-1. The limit of detection and limit of quantitation were found to be 0.033 μg mL-1and 0.1 μg mL-1 respectively. This method was tested and validated for various parameters according to ICH guidelines. The results demonstrated that the pro cedure is accurate, precise and reproducible (R.S.D. < 2 %). | 1 |
35371240 | INTRODUCTION R(+)-N-propargyl-1-aminoindan ("R-PAI") is also known as rasagiline and is a chiral compound with one asymmetric carbon atom in a five membered ring with an absolute (R) configuration which is produced as single enantiomer 1 .Rasagiline is a propargylamine-based drug indicated for the treatment of idiopathic Parkinson's disease 2 .In addition to rasagiline base, its acid addition salts (viz., mesylate, maleate, fumarate, tartrate, hydrobromide, p-acetate, benzoate, phosphate, tolunesulfonate and sulfate) are pharmaceutically acceptable 3 .Though different methods were reported for the determination of rasagiline mesylate in bulk and in pharmaceutical dosage forms [4][5][6][7] , only one each of RP-HPLC 8 and UV spectrophotometric method 9 were reported for the determination of rasagiline hemitartrate. In view of a lack of a commercial supplier and non development of visible spectrophotometric assay method till the date for rasagiline hemitartrate, the present study is aimed at its synthesis and development of a visible spectrophotometric analytical method for its bulk form by conducting systematic trials. | 2 |
35371240 | Absorption spectrum of coloured complex Into a 10 mL volumetric flask, 1 mL of 0.6% sulphanilic acid solution and 1 mL of 3 N hydrochloric acid solution were added.After cooling the contents to 4 0 C in an ice bath, 1 mL of 3% sodium nitrite was added and shaken for three minutes.To remove the added excess sodium nitrite, 1 mL of 4% sulphamic acid solution was added and shaken for two minutes.Then an aliquot of working standard drug solution (in the range 2 -10 mg) was added and the medium was made alkaline by the addition of 2 mL of 10% sodium hydroxide solution.The contents were made upto mark with distilled water to obtain yellow colored solution.The developed chromophore was scanned within the wavelength region of 400 -800 nm against a blank solution.The resulting spectrum was shown in Figure 1, and the absorption curve showed characteristic absorption maximum at 440 nm. | 6 |
35371240 | Theory of absorption spectrum of coloured complex Coupling of diazonium ion with drug molecules in basic medium to form azodyes has been used for the estimation of drugs by researchers 10,11 .In the proposed method, chromogen (diazonium salt) was prepared by diazotization of sulphanilic acid (having amino group) with sodium nitrite in Beer's Law Limit (Linearity, µg mL -1 ) 0 -10 3. Limit of quantitation (µg mL -1 ) 0.10 The absorption maximum for the coloured complex was found to be 440 nm.The absorption maxima of aqueous solutions of pure rasagiline and sulphanilic acid were 264 and 249 nm respectively.The l max of diazonium salt of sulphanilic acid was 270 nm.The blank solution (prepared on addition of sodium hydroxide to this diazonium salt of sulphanilic acid) didn't shift in the absorption maximum.However, a remarkable shift in the absorption maximum was observed when the blank solution was added to the drug solution.This was due to the formation of coupling product, which exhibits an absorption band peaking at 440 nm.This reaction was exploited to develop a spectrophotometric method for the determination of rasagiline hemitartrate in bulk drug form. | 7 |
35371240 | Mechanism of the color reaction The primary aromatic amines (p -nitro aniline / sulphanilic acid) react with NaNO 2 in acid medium in the temperature range of 0-3 0 C to give diazonium salt 12 .Colored azo compounds are formed by the coupling of the diazonium salts with strong nucleophiles (like electron rich aromatic / heteroaromatic compounds) 13 .The diazocoupling reaction can be considered as a proton eliminating condensation of a positive diazonium ion with another compound possessing an active hydrogen atom. The proposed method is based on the reaction of diazotization of amino group bearing sulphanilic acid (4-amino benzene sulphonic acid) with sodium nitrite in presence of hydrochloric acid at a temperature of 3 °C to form an electrophile, i.e., positive diazonium ion.The reaction is usually carried out in an ice bath and the excess sodium nitrite was removed by treatment with sulphamic acid.Since most of the diazonium salts are unstable 11 , the diazonium salt was used immediately. In alkaline medium, the positive diazonium ion couples at third position in rasagiline to form an yellow coloured azo product. | 8 |
35371240 | Optimization of reactions conditions Factors affecting the reaction conditions (concentrations of sulphanilic acid, hydrochloric acid, sodium nitrite, sulphamic acid and temperature) were studied by altering each variable in turn while keeping the others constant and the optimum conditions were established.The optimum conditions were selected based on their ability to give maximum absorbance and were maintained throughout the studies. In 1858, Peter Griess discovered the diazotization reaction 14 which requires three key components viz., an arylamine (Sulphanilic acid), a mineral acid (hydrochloric acid) and a source of nitrous acid (sodium nitrite) 15 . - As per the above equation, two equivalents of hydrochloric acid are required for sulphanilic acid.However, addition of excess acid is suggested to avoid the triazen formation.Triazen forms by the reaction of diazotized sulphanilic acid with the free sulphanilic acid 15 .Above 0.05N, with an increase in hydrochloric acid concentration, the rate of diazotization of sulphanilic acid increases 16 .Hence, the hydrochloric acid concentration was varied in between 0.05 to 1.5 N and the diazotization was found to be completed within three minutes for concentrations of 0.5 N and above up to 1.5 N. Hence, 1.0 N hydrochloric acid concentration was maintained in diazotization mixture by the addition of 1mL of 3 N HCl.Mixing of diazotization contents was done for six minutes though, the reaction completes within three minutes.The extended mixing is required because the reaction with nitrous acid is very slow towards the end of the diazotization 17 .To determine the optimum concentration of sulphanilic acid, the absorbance was studied by the addition of fixed volume (1 mL) of sulphanilic acid solution with variable concentrations (0.05-0.8%).Satisfactory results were obtained with 1 mL of a 0.6% sulphanilic acid solution. Carrying out the diazotization at low temperature is advantageous 18 due to (a) enhanced solubility of free nitrous acid which prevents the escape of nitrous gases from the acid medium and (b) improved stability of the diazotized sulphanilic acid (as the diazotized sulphanilic acid degrades to p-hydroxybenzene sulphonic acid at higher temperatures).Therefore, diazotization was carried out below 3 0 C. The effect of sodium nitrite concentration on diazotization was studied by the addition of 1 mL of NaNO 2 solutions with variation in the concentration range of 0.3 -5%.An increase in absorbance was observed with an increase in sodium nitrite concentration and became constant at 1%, above which, absorbance remained constant up to 5%.Therefore 1 mL of 3% sodium nitrite was chosen as an optimum value for the determination studies. Unlike hydrochloric acid, addition of excess sodium nitrite should be avoided due to destabilization of the diazotized salt by the surplus amount of nitrous acid produced in the medium 18 .Development of immediate blue colouration by moist potassium iodide starch paper helps the detection of surplus nitrous acid 17 .After completion of diazotization, the left over sodium nitrite can be destroyed by the addition of either urea or sulphamic acid 19 .These reagents convert the excess nitrite into nitrogen gas in acidic medium 20 .Sulphamic acid was preferred to remove the excess nitrite as it reacts faster than urea 21 . By knowing the leftover nitrite after diazotization, the amount of sulphamic acid to be added for its removal can be calculated from the stoichiometry of the respective destruction reaction.It was found to be 1 mL of 4% sulphamic acid. The effect of the addition of sodium hydroxide to the diazocoupling mixture was studied by following absorbance.The volumes of 10% sodium hydroxide varied from 1.0 to 3 mL.It was observed that maximum absorbance was observed by the addition of 2.0 ± 0.5 mL.A decrease in absorbance beyond 2.5 mL can be attributed to the partial decolorization of the dye at higher concentrations of alkali 22 .A decrease in colour intensity of the mixture at higher alkali conditions can be explained by consideration of acid-base equilibriums of diazonium compounds 23 .In general, aryldiazonium cations (e.g., phenyldiazonium) can lose their positive charge (i.e., high electrophilicity) and form diazohydroxides due to the attachment of anion to the terminal nitrogen atom.Further increase in alkalinity leads to subsequent deprotonation to form diazotates. Hence, in the present case, a maximum absorbance was observed when reaction mixture maintained at 0.5 M w.r.t.sodium hydroxide.In spite of rapid development of the yellow colored azo-dye, a maximum absorbance was attained after about 2 min at room temperature and the colour intensity was quite stable for at least one hour. | 9 |
35371240 | Comparison of validation parameters The results of validation parameters for the proposed method were within acceptable limits of ICH-2005 guidelines 25 .The correlation coefficient for the proposed method is 0.99937 indicating good linearity. % R S D i n r e c o ve r y o f r a s a g i l i n e hemitartrate by the proposed method (0.19-0.25) is lower compared to those values of visible spectrophotometric methods proposed by other workers 4,26 for determination of its equivalent drug -rasagiline mesylate.LOD and LOQ values of the proposed method for rasagiline hemitartrate (0.033 and 0.1) are lower compared to those values of visible spectrophotometric methods proposed by other workers 4,26 for determination of its equivalent drug -rasagiline mesylate.Hence, the proposed diazotization method for visible spectrophotometric determination of Rasagiline hemitartrate was proved to be better compared to red-ox / ion-pair complex formation methods proposed for the determination of its equivalent drug -rasagiline mesylate. | 16 |
220843287 | Neodymium complexes containing N-heterocyclic carbene (NHC) ligands, NdCl3[1,3-R2(NCH=)2C:]·THFx(Nd1: R = 2,6-iPr2C6H3, x = 0; Nd2: R = 2,6-Et2C6H3, x = 1; Nd3: R = 2,4,6-Me3C6H2, x = 1) were synthesized and employed as precatalysts for the coordination polymerization of conjugated dienes (butadiene and isoprene). In combination with triisobutylaluminium (TIBA), Nd1 promoted butadiene polymerization to produce extremely high cis-1,4 (up to 99.0%) polybutadienes with high molecular weight (Mw = 250–780 kg·mol−1). The Nd1/TIBA catalytic system also exhibited both high catalytic activity and cis-1,4 selectivity (up to 97.8%) for isoprene polymerization. The catalytic activity, molecular weight and molecular weight distribution of resulting polydienes were directly influenced by Al/Nd molar ratio, aging method, and polymerization temperature. Very interestingly, the high cis-1,4 selectivity of the catalyst towards butadiene and isoprene kept almost unchanged under different reaction conditions. The cis-1,4 polyisoprenes with high molecular weight (Mw = 210–530 kg·mol−1) and narrow molecular weight distribution (Mw/Mn = 1.9–2.7) as well as high cis-1,4 selectivity (~97%) could be synthesized by using the aged Nd1/TIBA catalytic system in the presence of isoprene (100 equivalent to Nd) at low Al/Nd molar ratios of 6–10. Polyisoprenes with low molecular weights (Mw = 12–76 kg·mol−1) and narrow molecular weight distributions (Mw/Mn = 1.7–2.6) were obtained by using Nd2 and Nd3 as precatalysts, indicating that the molecular weight of resulting polyisoprenes can be adjusted by changing the substitutes of ligand in Nd complex. | 1 |
220843287 | INTRODUCTION Cis-1,4 selective polymerization of conjugated dienes, e.g. butadiene and isoprene, is of great importance in synthetic rubber industry to produce the high cis-1,4 polydienes with excellent properties such as excellent elasticity, high fatigue and crack resistance. [1] Catalyst, which mainly decides the catalytic activity and the microstructure of resulting polymers, plays an important role in the industrial production of polydienes. [2−5] Therefore, much effort has been devoted to developing various catalysts for producing polydienes with high cis-1,4-regularity and controlled molecular weight. [2−5] Rare earth based catalysts stand out as being highly active and selective for butadiene and isoprene polymerizations. [2−5] In general, these rare earth based catalysts are divided into two types: Ziegler-Natta catalytic system and cationic catalytic system. [3−5] The Ziegler-Natta rare earth metal catalysts, mainly the binary systems (LnCl 3 -R 3 Al) and ternary systems (LnL 3 -R 3 Al-X, Ln = lanthanide; L = carboxylate, phosphate, alkyl, or aryl oxide; X = compounds containing halogen atom), have been used in the synthetic rubber industry because of their advantages in easy preparation, thermal stability, and low moisture and air sensitivity. [3−5] The addition of oxygen-containing ligands, e.g. alcohols [6−8] and tetrahydrofuran, [9] enhanced the catalytic activity of the binary systems. Bidentate amidinate, [10−12] β-diketimines, [13] iminopyrrole, [14] indolide-imine, [15] aminopyridinato, [16] aminoindolyl, [17] alkoxy N-heterocyclic carbene [18] ligands, and tridentate pincer ligands, such as N^C^N, [19] C^C^C, [20] P^N^P, [22,23] N^C^O, [24] N^C^S, [24] N^N^O, [25,26] N^N^N [27−29] ligands were also used in the preparation of lanthanide complexes. Activated by organoborate cocatalyst, the obtained cationic catalytic systems display high catalytic activity and cis-1,4 selectivity toward conjugated diene polymerization. [10−29] The chemical structures of ancillary ligands could steer the behavior of the coordination polymerization and characteristics of the resulting polymers. The concept that the ligand plays regulatory role in the catalytic behavior of the catalyst was used to design Ziegler-Natta rare-earth metal catalysts. In the presence of triisobutylaluminium (Al i Bu 3 ), neodymium complexes containing heterocyclic Shiff base, [30] 8hydroxyquinolines, [31] quinolinylcarboxylates, [32] or NCN-pincer ligand [33,34] show high catalytic activity for isoprene polymerization with high cis-1,4 stereospecificity (95%−98%). Nheterocyclic carbene (NHC) has become an organo-catalyst and ubiquitous ligand in organometallic chemistry because of its extraordinary electron richness and facile access to structurally diverse analogues. [35] Scandium trialkyl complexes containing N-heterocyclic carbene ligand have been reported as precatalysts for α-olefin polymerization with excellent catalytic activity. [36,37] We also reported that the copolymerization of ethylene with propylene was realized by vanadium complexes containing NHC ligands and both the catalytic activity and microstructure of the resulting copolymers were influenced by the chemical structure of the NHC ligands. [38,39] Therefore, introduction of NHC ligand to NdCl 3 is of great interest for the development of a novel neodymium-based catalytic system with both high activity and regioselectivity for the coordination polymerization of the conjugated dienes. Herein, the synthesis of novel NdCl 3 ·NHC·THFx complexes (NHC: 1,3-R 2 (NCH=) 2 C:; Nd1: R = 2,6-i Pr 2 C 6 H 3 , x = 0; Nd2: R = 2,6-Et 2 C 6 H 3 , x = 1; Nd3: R = 2,4,6-Me 3 C 6 H 2 , x = 1) and their catalytic behavior for the coordination polymerizations of butadiene and isoprene upon activation with Al i Bu 3 were investigated. The Nd1-based catalytic system showed both high catalytic activity and 1,4-selectivity for conjugated diene polymerizations, affording polybutadienes with extremely high cis-1,4 content up to 99.0% and polyisoprenes with high cis-1,4 content of 97.8% as well as high molecular weight and narrow molecular weight distribution. | 2 |
220843287 | EXPERIMENTAL General Considerations All manipulations of air-and moisture-sensitive compounds were performed in a nitrogen atmosphere using standard Schlenk techniques or under a nitrogen atmosphere in a drybox. Tetrahydrofuran (THF, Beijing Chemical Works) was distilled under nitrogen atmosphere and refluxed over sodium benzophenone for dehydration, and then stored in the drybox in the presence of molecular sieves (4Å). NdCl 3 ·xTHF [9] and NHC ligand [40] were prepared according to the reported methods. Chlorobenzene (C 6 H 5 Cl, Tianjin Fuchen Chemical Co.) was freshly distilled from phosphoric anhydride. Hexanes and cyclohexane (Beijing Yanshan Petrochemical Co.) were dried over calcium hydride (CaH 2 ) and distilled before use. Isoprene (purity: 99.9%, Beijing Yanshan Petrochemical Co.) was freshly distilled from CaH 2 before use. Butadiene (Beijing Yanshan Petrochemical Co.) and triisobutylaluminium solution in hexanes (0.74 mol·L −1 , Beijing Yanshan Petrochemical Co.) were used as received. | 3 |
220843287 | Procedure of Conjugated Diene Polymerization All the operations were conducted under an atmosphere of dry nitrogen. For the polymerization using the in situ prepared catalyst, the conjugated diene monomers (butadiene or isoprene) and solvent were introduced into a vessel and Al i Bu 3 was added. Then, the solution of Nd complex was introduced into the vessel to start the coordination polymerization of conjugated diene at a defined temperature. For the polymerization using the aged catalyst, the mixture of Nd complex and Al i Bu 3 in the presence of different amounts of monomer was aged at the fixed temperature for the designated time in advance. The conjugated diene monomers (butadiene or isoprene) and solvent were introduced into a vessel and then the aged catalyst solution was added to start the coordination polymerization of conjugated diene at a defined temperature. The vessel with stirring was placed in a bath with constant temperature during the polymerization. After a definite time, the polymerization was terminated by addition of ethanol containing 1% of 2,6-di-tertbutyl-4-methylphenol. Then the mixture was poured into ethanol containing a small amount of hydrochloric acid. The precipitated polymer was further washed by ethanol and then was dried under vacuum at 45 °C until a constant weight. | 4 |
220843287 | Characterization of Resulting Polymers Molecular weights of resulting polybutadienes and polyisoprenes, i.e. number-average molecular weight (M n ), weightaverage molecular weight (M w ), and polydispersity index (PDI, M w /M n ), were determined by gel permeation chromatography (GPC) using a Waters 1515-2410 system equipped with Waters RI 2410 and UV 2489 detectors and four Waters styragel HT3-4-5-6 columns (Milford, MA). The polymer sample was dissolved in THF with concentration of 2 g·L −1 . THF was used as eluent and the flow rate of the mobile phase was 1.0 mL·min −1 at 30 °C. The calibration curve was obtained by polystyrene standard. The contents of cis-1,4, trans-1,4, and 1,2 structures of resulting polydienes were determined using FTIR analysis according to the reported method. [42] The film of the copolymer was prepared by spreading a small amount of dichloromethane (CH 2 Cl 2 ) solution of the copolymer on the slice of KBr after the evaporation of CH 2 Cl 2 . The copolymer was characterized on a Nexus 670 FTIR spectrophotometer (Nicolet, Medison, WI). | 5 |
220843287 | Synthesis of Nd Complexes with NHC Ligands The reaction of equimolar quantities of NHC ligands and NdCl 3 ·xTHF (x = 1,2,3) in THF under nitrogen at 25 °C for 5 h afforded the Nd complexes Nd1−Nd3, as shown in Scheme 1. All the paramagnetic complexes Nd1−Nd3 were characterized by elemental analysis and the Nd contents of these complexes were determined by titration. The results indicated that one THF molecule was incorporated in the complexes of Nd2 and Nd3, respectively. Comparably, no THF molecule existed in Nd1 complex due to the bulky isopropyl substitutes on phenyl rings in ligand. | 6 |
220843287 | catalytic system The neodymium complex Nd1 containing NHC ligand with bulky isopropyl substituents at the ortho positions of the phenyl rings was employed as precatalysts and triisobutylaluminum (Al i Bu 3 , Al) was used as a cocatalyst to investigate the coordination polymerization of conjugated dienes (butadiene and isoprene). Butadiene polymerizations and isoprene polymerizations under various Al/Nd molar ratios, polymerization temperatures (T p ), and polymerization time (t p ) were investigated using prepared Nd1/Al i Bu 3 catalytic system, in which the active centers formed in situ in the polymerization system. The experimental results are summarized in Table 1. It can be seen from the data in Table 1 that the conversion of butadiene and catalytic activity increased along with an increase in Al/Nd molar (entries 1−3 and 5−8). The neodymium complex Nd1 displayed good catalytic activity (2.8 × 10 4 g·mol −1 of Nd) for butadiene polymerization at Al/Nd molar ratio of 50 (entry 4 in Table 1), albeit poor catalytic activities were observed at low Al/Nd molar ratio (entries 1 and 2 in Table 1). Remarkably, polybutadiene with high cis-1,4 content of ~99.0% and high molecular weight (M w = 540 kg·mol −1 ) was obtained. The conversion of butadiene could be improved obviously from 6% to 20% at the Al/Nd ratio of 15 by increasing T p from 25 °C to 50 °C. As Al/Nd molar ratio increased from 15 to 50, the conversion of butadiene increased from 20% to 60%, and the catalytic activity increased from 1.4 × 10 4 g·mol −1 of Nd to 4.1 × 10 4 g·mol −1 of Nd (entries 5−8 in Table 1). Polybutadiene with high molecular weight (M w = 470 kg·mol −1 ) and uniform molecular weight distribution was afforded at the Al/Nd molar ratio of 15 (entry 5 in Table 1). However, the molecular weight distribution became broader (4.6−14.0) indicating that multiple active species formed or chain transfer reaction speeded up with increasing Al/Nd molar ratio. Interestingly, the distinguished cis-1,4 selectivity kept almost unchanged (97.9%−98.8%) in a broad range of Al/Nd molar ratio from 15 to 50 (entries 5−8 in Table 1). Overall, polybutadienes with high cis-1,4 contents were obtained by polymerization of butadiene using Nd1/Al i Bu 3 catalytic system. The Al/Nd molar ratio and polymerization temperature have an obvious influence on the catalytic activity, molecular weight, and molecular weight distribution. | 8 |
220843287 | Isoprene polymerization with the unaged Nd1/Al i Bu 3 catalytic system According to the above investigation on butadiene polymeri- zation, Nd1 containing NHC ligand with bulky isopropyl substituents at the ortho positions of the phenyl rings was also selected as precatalyst for the coordination polymerization of isoprene herein. The effects of Al/Nd molar ratio and polymerization temperature on isoprene polymerization were investigated using unaged Nd1/Al i Bu 3 catalytic system. The results are summarized in Table 1 (entries 9−20). The isoprene polymerization was carried out and the yield of polymer was negligible under the similar polymerization conditions to those for butadiene polymerization. Negligible polyisoprene was obtained in the mixed solvent of hexane and cyclohexane, possibly due to the poor solubility of catalyst in the polymerization system. Chlorobenzene was firstly selected as a good solvent in the polymerization of isoprene to investigate systematically the effects of chemical structure of ligands, preparation process of catalytic system, and polymerization conditions on the catalytic activity and the microstructure of the resulting polymers. The amount of cocatalyst, which is usually expressed by the molar ratio of Al/Nd, has a significant influence on the catalytic activity and molecular weight and molecular weight distribution of the resulting polyisoprenes. It can be seen from Table 1 that an increase in isoprene conversion and catalytic activity could be noticed as the Al/Nd molar ratio increased (entries 10−14). The isoprene conversion of 71% and the catalytic activity of 5.0 × 10 4 g·mol −1 of Nd at T p of 50 °C could be obtained at Al/Nd molar ratio of 30 (entry 14 in Table 1). The molecular weight of the resulting polyisoprene decreased with an increase in Al/Nd molar ratio probably due to the more chain transfer reaction to Al i Bu 3 at higher Al/Nd molar ratio. It is worth noting that the microstructure of the resulting polyisoprenes was not affected by the change of Al/Nd molar ratio. As shown in Table 1, polyisoprenes with cis-1,4 content of ca. 96% could be prepared at T p of 50 °C when the Al/Nd molar ratio increased from 10 to 30. Isoprene polymerizations were carried out at polymerization temperature (T p ) ranging from 30 °C to 60 °C and the results are given in Table 1 (entries 15−20). It can be clearly observed that T p influenced the isoprene conversion, catalytic activity, and molecular weight, molecular weight distribution, and cis-1,4 content of the resulting polyisoprenes. It can be seen from Table 1 that isoprene conversion greatly increased from 50% to 84% and catalytic activity increased from 3.5 × 10 4 g·mol −1 of Nd to 5.9 × 10 4 g·mol −1 of Nd when T p was elevated from 30 °C to 50 °C (entries 15−20). However, the overall catalytic activity and isoprene conversion decreased when T p was higher than 50 °C since the catalyst deactivation became more prominent at higher polymerization temperature. Similar to other reported catalytic systems, [30,31,33] a slight decrease in cis-1,4 content in polymer products with increasing polymerization temperature can be observed. The GPC traces of the resulting polyisoprenes prepared at different temperatures from 30 °C to 50 °C are displayed in Fig. 1. It can be seen that all the GPC traces of the resulting polyisoprenes exhibit bimodal and broad molecular weight distribution. The overall molecular weight decreased greatly and the molecular weight distribution became broad with an increase in T p , as shown in Fig. 2. The chain transfer side reaction could be accelerated with increasing polymerization temperature and thus the overall molecular weight decreased greatly. | 9 |
220843287 | Isoprene polymerization using the aged Nd1/Al i Bu 3 catalytic system The catalyst components of Nd1 and Al i Bu 3 reacted to form the active centers prior to the addition to the monomer solution, which is also referred to as catalyst aging process. Both the aging temperature (T a ) and aging time (t a ) played important roles in the formation of active centers in the aging process of catalyst. The obtained catalyst solution after the aging process was used for isoprene polymerization. The experimental results of isoprene polymerization using the above aged catalyst are displayed in Table 2 (entries 2−7) and isoprene polymerization using the unaged catalyst is also displayed in Table 2 (entry 1) for comparison. It can be found from Table 2 that isoprene conversion increased greatly from 2% to 25%−95% and catalytic activity increased greatly from 0.2 × 10 4 g·mol −1 of Nd to 6.7 × 10 4 g·mol −1 of Nd by using the aged catalyst instead of unaged catalyst under similar polymerization conditions. The catalytic behavior of aged Nd1/Al i Bu 3 catalytic system is affected by T a . The isoprene conversion increased from 25% to 87% and the catalytic activity increased from 1.8 × 10 4 g·mol −1 of Nd to 6.1 × 10 4 g·mol −1 of Nd along with an increase in T a from 40 °C to 60 °C at t a of 30 min, while the cis-1,4 selectivity kept at around 96.5% (entries 2, 4, and 7 in Table 2). The isoprene conversion and catalytic activity increased while the Table 2). All the results indicate that the catalytic activity could be remarkably improved by aging process of the catalysts. The reaction of Nd complex with Al i Bu 3 results in the formation of Nd compounds with σ-alkyl bonds in the absence of monomer. However, the reaction of Nd complex with Al i Bu 3 results in the formation of the π-allyl Nd complexes in the presence of monomer, which exhibit a higher stability than that of Nd compounds with σ-alkyl bonds. The isoprene polymerization using the aged Nd1/Al i Bu 3 catalyst in the presence of isoprene (Ip/Nd = 100) prepared for different aging time (t a ) was further investigated and the experimental results are displayed in Table 2 (entries 8−12). In order to distinguish two different aging methods and express clearly, aging method without isoprene is expressed as method A, while aging method with isoprene is expressed as method B. Isoprene conversion in the polymerization process could reach 99% and the catalytic activity could reach 6.9 × 10 4 g·mol −1 of Nd even at T p of 0 °C for polymerization time of 14 h by using the aged ternary catalyst with t a of 3 min. A very high conversion of 93% and catalytic activity of 6.5 × 10 4 g·mol −1 of Nd can also be obtained with the aging time of 9 min, which implies enough operation time. However, monomer conversion decreased to 21% if t a was 60 min, which was a different trend from that in aging method A. The molecular weight of the obtained polyisoprenes was also affected by aging time. The molecular weight of polyisoprenes increased with an increase in aging time, which might be attributed to the decreasing amount of active species in the catalytic system with increased t a . The aging time hardly affected the cis-1,4 content of the resulting polyisoprenes, indicating that the catalytic system displayed high cis-1,4 selectivity at even at long aging time. Although high isoprene conversion and preparation of polyisoprene with high molecular weight were realized, the molecular weight distribution was still broad. Therefore, the isoprene polymerizations with ternary catalyst (B) with low Al/Nd molar ratios were further conducted at low T p of −15 °C. As shown in Table 2 (entries 13−22), the molecular weight distribution of resulting polyisoprenes at low T p of −15 °C became much narrower than those of polyisoprenes synthesized at T p s of 0 and 25 °C, although the conversion of isoprene and catalytic activity decreased to 21%−53% and 1.5 × 10 4 − 3.7 × 10 4 g·mol −1 of Nd, respectively. The isoprene conversion of 53% could be obtained even the Al/Nd molar ratio was decreased to 8 by optimization of t a (entry 17 in Table 2). The regular effect of Al/Nd molar ratio on isoprene conversion was not observed. Very importantly, polyisoprenes with high molecular weight (M w ) ranging from 210 kg·mol −1 to 530 kg·mol −1 and narrow molecular weight distribution (M w /M n = 1.9−2.7) could be obtained at various Al/Nd molar ratios and t a s at low T p of −15 °C (entries 13−22 in Table 2). The relatively unimodal GPC traces of resulting polyisoprenes are displayed in Fig. 3. The influences of t a on M w and M w /M n were different at various Al/Nd molar ratios due to the complicated reaction of Nd1 with Al i Bu 3 in the presence of isoprene. Polyisoprene with high molecular weight (530 kg·mol −1 ) and narrow molecular weight distribution (M w /M n = 2.4) could be successfully synthesized at T p of −15 °C using the ternary catalyst (Ip/Al/Nd molar ratio = 100/6/1) by aging method B. Moreover, higher cis-1,4 selectivity (96.9%−97.6%) was observed using the ternary catalyst than that using unaged binary catalyst (entries 8−22 in Table 2 versus entries 9−19 in Table 1). The representative FTIR spectra of resulting polyisoprenes prepared by using aged ternary catalyst and unaged binary catalyst are shown in [27][28][29][30][31][32]42] As shown in Fig. 4, a stronger band at 1128 cm −1 and a weak band at 889 cm −1 can be observed in the FTIR spectrum of polyisoprene prepared by the aged ternary catalyst as compared with that of polyisoprene prepared by the unaged binary catalyst, indicating that the aged ternary catalyst displayed higher cis-1,4 selectivity than that of the unaged binary catalyst. The results of isoprene polymerization using aged catalyst indicate that the catalytic activity could be improved obviously by aged catalyst. Polyisoprene with high molecular weight and broad molecular weight distribution could be afforded by using aged Nd1/Al i Bu 3 catalytic system, while polyisoprene with high molecular weight and narrow molecular weight distribution could be afforded by using aged ternary catalyst (Ip/Nd1/Al i Bu 3 ). | 10 |
220843287 | Effect of Ligands in Nd Complexes on Catalytic Activity and Microstructure of Resulting Polydienes Isoprene polymerizations by using Nd complexes containing NHC ligands with ethyl (Nd2) or methyl substitutes (Nd3) at the N-aryl ring were investigated. The experimental results of isoprene polymerizations at various Al/Nd molar ratios are summarized in Table 3. The aged ternary catalysts (Ip/Nd2/ Al i Bu 3 and Ip/Nd3/Al i Bu 3 ) prepared by aging method B exhibited both good activity and high cis-1,4 selectivity at relatively high Al/Nd molar ratios. At optimized Al/Nd molar ratio, the isoprene conversion and catalytic activity for Nd2 were 61% and 4.4 × 10 4 g·mol −1 of Nd, respectively. Meanwhile, the isoprene conversion and catalytic activity for Nd3 were 83% and 6.0 × 10 4 g·mol −1 of Nd, respectively. Polyisoprenes prepared by using precatalyst Nd2 at the Al/Nd ratios of 15 and 20 exhibited high cis-1,4 content of 97.8% (entries 1 and 2 in Table 3). The molecular weight and molecular weight distribution of the resulting polymers were significantly influenced by the structure of the Nd complex. Compared to polyisoprenes prepared with Nd1, polyisoprenes with the drastically lower molecular weight (M w = 12−51 kg·mol −1 for Nd2 and 15−76 kg·mol −1 for Nd3) and unimodal molecular weight distribution (M w /M n = 1.7−2.6) were afforded by using Nd2 or Nd3 as the precatalyst (entries 1−7 in Table 3). The result suggests that a uniform active species existed during polymerization of isoprene using Nd2 or Nd3 as precatalyst. 26 28 30 32 34 36 Elution time (min) Al/Nd = 6, t a = 3 min (entry 20 in Table 2) Al/Nd = 8, t a = 3 min (entry 17 in Table 2) Al/Nd = 10, t a = 3 min (entry 13 in Table 2) Signal intensity Table 2). | 11 |
220843287 | Fig. 4 Representative FTIR spectra of resulting polyisoprenes prepared by using aged ternary catalyst (entry 13 in Table 2) and unaged binary catalyst (entry 17 in Table 1). The average number of polymer chains (n calcd ) can be calculated by the ratio of m p and M n due to the narrower molecular weight distribution, where m p is the weight of the resulting polyisoprene and M n is number-average molecular weight of the resulting polyisoprene (kg·mol −1 ). The theoretical numbers of polymer chains (n theo ) in the copolymerization system were 4.0 × 10 −5 mol. It can be observed that n calcd is much higher than n theo for Nd2/Al i Bu 3 catalytic system (Al/Nd = 20 and 30) and Nd3/Al i Bu 3 catalytic system (Al/Nd = 30 and 40), which is attributed to serious chain-transfer reaction to a cocatalyst during isoprene polymerization. The molecular weight and molecular weight distribution of resulting polyisoprenes are greatly affected by the structure of the ligand (as shown in Fig. 5). Polyisoprenes with the low molecular weights and narrow molecular weight distributions were obtained by using complexes Nd2 and Nd3 bearing NHC ligands with ethyl or methyl substitutes at the N-aryl ring due to the combination of steric hindrance effect and electronic effect of the ligands. Comparatively, polyisoprenes with the higher molecular weights were prepared by using complex Nd1 containing NHC ligand with bulky isopropyl substitutes. Therefore, polyisoprenes with low or high molecular weight and narrow molecular weight distribution could be afforded by changing the substitutes at the N-aryl rings of the Nd complex. The effect of ligand on the molecular weight of resulting polyisoprenes also indicates that the NHC ligand was associated with the active Nd centers during polymerization of isoprene. | 12 |
54677822 | The oxidative chemical polymerization of o-phenylenediamine (OPDA) was studied in hydrochloric acid solution using potassium dichromate as oxidant at 5C. The effects of potassium dichromate, hydrochloric acid, and monomer concentrations on the polymerization reaction were investigated. The order of reaction with respect to potassium dichromate, hydrochloric acid, and monomer concentration was found to be 1.011, 0.954, and 1.045, respectively. Also, the effect of temperature on the polymerization rate was studied and the apparent activation energy of the polymerization reaction was found to be 63.658 kJ/mol. The obtained polymer was characterized using XPS, IR, UV-visible, and elemental analysis. The surface morphology of the obtained polymers was characterized by X-ray diffraction and transmission electron microscopy (TEM). The TGA analysis was used to confirm the proposed structure and number of water molecules in each polymeric chain unit. The ac conductivity (σac) of (POPDA) was investigated as a function of frequency and temperature. The ac conductivity was interpreted as a power law of frequency. The frequency exponent (s) was found to be less than unity and decreased with the increase of temperature, which confirms that the correlated barrier hopping model was the dominant charge transport mechanism. | 1 |
54677822 | Introduction Polyaniline as an electrically conductive polymer has attracted considerable attention, because of its excellent environmental stability in the electroconducting form and electrical and optical properties [1,2].It has various potential applications in many high performance devices [3][4][5][6][7][8][9].A common feature of conducting polymer is conjugation of -electrons extending over the length of the polymer backbone [10].Polymerization of conducting polymer may be performed by chemical [11] or electrochemical [12] methods.Various chemical oxidizing agents, such as potassium dichromate [13], potassium iodate [14], hydrogen peroxide [15], ferric chloride, or ammonium persulphate, were used [16].The applications of polyaniline are limited due to its poor processability [17], which is true for most conducting polymers.Several studies have been done in order to improve the solubility of polyaniline; among them is using functionalized protonic acids as dopant, like p-toluenesulphonic acid, octyl-benzene-sulphonic acid, dodecyl benzene-sulphonic acid [18], poly(styrene) sulphonic acid [19], and phosphoric acid esters [20].An alternative method to obtain soluble conductive polymers is the polymerization of aniline derivatives.The studied aniline derivatives are alkyloxy, hydroxy, and chloroaniline and substitution at the nitrogen atom was reported by Sayyah et al. [21][22][23][24] to improve the solubility of polyaniline.The substituted group of aniline affects not only the polymerization reaction but also the properties of the polymer obtained.The kinetics of chemical polymerization of 3-methylaniline, 3-chloroaniline, 3-hydroxyaniline, 3-methoxyaniline, and N-methyl aniline in hydrochloric acid solution using sodium dichromate as oxidant and characterization of the polymer obtained by IR, UV-visible and elemental analysis, X-ray diffraction, 2 International Journal of Polymer Science scanning electron microscopy, TGA-DTA analysis, and ac conductivity have been investigated by Sayyah et al. [25][26][27][28]. The present work intends to study the kinetics of the oxidative chemical polymerization of o-phenylenediamine in aqueous HCl medium and potassium dichromate as oxidant.The obtained polymer is characterized by XPS, IR, UVvisible, TGA, elemental analysis, X-ray, transmission electron microscopy (TEM), and ac conductivity measurements. | 2 |
54677822 | Oxidative Aqueous Polymerization of O-Phenylenediamine Monomer. The polymerization reaction was carried out in a well-stoppered conical flask of 250 mL capacity; addition of OPDA amount in 25 mL HCl of known molarity was followed by the addition of the required amount of potassium dichromate as oxidant in water (25 mL) to the reaction mixture.The orders of addition of substances were kept constant in all the performed experiments.The stoppered conical flasks were then placed in an automatically controlled thermostat at the required temperature.The flasks were shaken (15 shakings/10 s/15 min) by using an automatic shaker.The flasks were filtrated using a Buchner funnel; then the obtained polymer was washed with distilled water and finally dried till constant weight in vacuum oven at 60 ∘ C. | 4 |
54677822 | Elemental Analysis, Infrared, and Ultraviolet Spectroscopy. The carbon, hydrogen, and nitrogen contents of the prepared polymer were carried out in the microanalytical laboratory at Cairo University by using oxygen flask combustion and a dosimat E415 titrator (Switzerland). The infrared spectroscopic analysis of the prepared polymer was carried out in the microanalytical laboratory at Cairo University by using a Shimadzu FTIR-430 Jasco spectrophotometer and KBr disc technique. The ultraviolet-visible absorption spectra of the monomer and the prepared polymer sample were measured using Shimadzu UV spectrophotometer (M 160 PC) at room temperature in the range 200-400 nm using dimethylformamide as a solvent and reference. | 5 |
54677822 | X-Ray Photoelectron Spectroscopy (XPS). An XPS spectrum was obtained on XPS-thermo scientific spectrometer, Model: K-ALDH in Central metallurgical research and development institute (CMRDI).Polymer was mounted on a standard sample holder using double-sided adhesive tape.Survey and XPS spectra were obtained with Al K monochromatic X-ray with the resolution of 0.7 eV. | 6 |
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