Molecular simulation, characteristics and mechanism of thermal-responsive acetylated amylose V-type helical complexes.

To explore the thermal-responsive characteristics of acetylated amylose-guest V-type helical complexes (AAGHCs) and their potential use as thermal-responsive drug carriers, different types of AAGHCs were built, in which acetylated amylose was used as a host, and iodine, propofol, or hexane was utilized as the guest molecule. Their thermal-responsive characteristics were investigated through molecular dynamic (MD) simulation and corresponding experiments. MD simulation showed that the thermal-responsive helix-unfolding and guest-release behavior in AAGHCs, and the complete unfolding of AAGHC could be divided into brewing, triggering and collapsing periods. Energy analysis revealed that the Lana-Jones potential is an important binding energy that bridges host and guest molecules and enhances the stability of the helix. The various types or number of guests showed different binding energies. The stronger the binding energy, higher is the temperature required to trigger the unfolding of the helix and the releasing of guests. FT-IR and X-ray diffraction analyses confirmed the structures of AAGHCs. The change in hydrated size, and UV-VIS absorption of AAGHCs at high temperatures both confirmed the thermal-responsiveness of AAGHCs. The fluorescence fluctuation of loaded 7-hydroxycoumarin reflected the same thermal-responsive process and mechanism as MD simulation. This study provides meaningful theoretical guidance for the design of thermal-responsive drug carriers based on acetylated amylose-guest V-type helical complexes.

Engineering of Biocompatible Coacervate-Based Synthetic Cells.

Polymer-stabilized complex coacervate microdroplets have emerged as a robust platform for synthetic cell research. Their unique core-shell properties enable the sequestration of high concentrations of biologically relevant macromolecules and their subsequent release through the semipermeable membrane. These unique properties render the synthetic cell platform highly suitable for a range of biomedical applications, as long as its biocompatibility upon interaction with biological cells is ensured. The purpose of this study is to investigate how the structure and formulation of these coacervate-based synthetic cells impact the viability of several different cell lines. Through careful examination of the individual synthetic cell components, it became evident that the presence of free polycation and membrane-forming polymer had to be prevented to ensure cell viability. After closely examining the structure-toxicity relationship, a set of conditions could be found whereby no detrimental effects were observed, when the artificial cells were cocultured with RAW264.7 cells. This opens up a range of possibilities to use this modular system for biomedical applications and creates design rules for the next generation of coacervate-based, biomedically relevant particles.

Improving ordered arrangement of the short-chain amylose-lipid complex by narrowing molecular weight distribution of short-chain amylose.

To investigate effect of molecular weight distribution on formation of the short-chain amylose-lipid complex, debranched waxy rice starch (Unf) was fractionated into F1, F2 and F3, and the four fractions were used to complex with palmitic acid (C16:0). The peak molecular weight was in the order of F1 < Unf < F2 < F3, and the distribution was in the order of Unf> F3 > F1 > F2. XRD and DSC analysis indicated that the Unf-C16:0 complex was amorphous with the melting temperature (Tm) < 100 °C while the F3-C16:0 complex was highly ordered with Tm > 100 °C. Moreover, the F1-C16:0 complex was more ordered than the Unf-C16:0 complex, although F1 had lower molecular weight. These results suggested that improving homogeneity of short-chain amylose promoted ordered arrangement of the short-chain amylose-lipid complex. Moreover, the F3-C16:0 complex was the most resistant, which accorded with its highest crystallinity.

Chitosan coating of zein-carboxymethylated short-chain amylose nanocomposites improves oral bioavailability of insulin in vitro and in vivo.

Non-invasive means of insulin administration circumvent some of the inconveniences of injections. Oral administration in particular is convenient, pain-free, and allows favorable glucose homeostasis, but is subject to chemical instability, enzymatic degradation, and poor gastrointestinal absorption. Natural polymeric nanoparticles have emerged as a promising oral delivery system for peptide therapeutics due their safety, biocompatibility, and stability. In this study, self-assembled nanocomposites from chitosan (CS) and insulin-loaded, zein-carboxymethylated short-chain amylose (IN-Z-CSA) nanocomposites were synthesized to improve oral bioavailability of insulin. The optimized IN-Z-CSA/CS0.2% nanocomposites exhibited an average size of 311.32±6.98 nm, a low polydispersity index (0.227±0.01), a negative zeta potential (43.77±1.36 mV), an encapsulation efficiency of 89.6±0.9%, and a loading capacity of 6.8±0.4%. The IN-Z-CSA/CS0.2% nanocomposites were stable in storage conditions. The transepithelial permeability of the N-Z-CSA/CS0.2% nanocomposites was 12-fold higher than that of insulin. Cellular uptake studies revealed that the IN-Z-CSA/CS0.2% nanocomposites were internalized into Caco-2 cells by both endocytosis and a paracellular route. Additionally, in pharmacological studies, orally administered IN-Z-CSA/CS0.2% nanocomposites had a stronger hypoglycemic effect with a relative bioavailability of 15.19% compared with that of IN-Z-CSA1.0% nanocomposites. Furthermore, cell toxicity and in vivo tests revealed that the IN-Z-CSA/CS0.2% nanocomposites were biocompatible. Overall, these results indicate that the IN-Z-CSA/CS0.2% nanocomposites can improve oral bioavailability of insulin and are a promising delivery system for insulin or other peptide/protein drugs.

Low-density graphitic films prepared from iodine-doped enzymatically synthesized amylose films as carbonization precursors.

We have developed a novel approach for preparing low-density graphitic films using iodine-doped enzymatically synthesized amyloses (ESAs) with strictly controlled molecular weights as carbonization precursors. All of the iodine-doped ESA films retained their film structures and morphologies, even after the heat-treatment at 800 °C and 2600 °C. Therefore, iodine doping plays an indispensable role in retaining film structure and morphology during the carbonization of ESA polysaccharides. It was also elucidated that the carbonization yields of the ESA films can be controlled by changing the conditions of iodine doping process. The bulk densities of the graphitic films are varied from 0.08 to 0.42 g/cm3 dependent on the doping level. In addition, the capacitance performances of the graphite films prepared from the ESAs are investigated using cyclic voltammetry and galvanostatic charge/discharge procedures. The potential utility of the carbonized and graphitized ESA films for supercapacitors was revealed. This approach may broaden the application and even the swill processing of polysaccharides.

Biosynthesis of superparamagnetic polymer microbeads via simple precipitation of enzymatically synthesized short-chain amylose.

Here, we report a simple and non-emulsion based approach to prepare starch-based magnetic polymer beads with well-defined size, shape and dispersibility in aqueous environment through co-precipitation of enzymatically synthesized short chain amylose (SCA) and dextran-coated iron oxide nanoparticles (Dex@IONPs). The size and morphology of magnetic polymer beads (MPBs) were controlled by employing Dex@IONPs as a seeding agent. The resulting superparamagnetic amylose microbeads (SAMBs) were readily functionalized with antibody through one step reaction using a linker protein, which showed great capture efficiency (>90%) and specificity for target bacteria present in complicated food matrix. The excellent magnetic sensitivity also enabled the SAMBs readily assembled into ordered 1D arrays by external magnetic field whose structure could permanently be fixed by SCA-mediated precipitation process.

Syntheses and structure-membrane active antimicrobial activity relationship of alkylamino-modified glucose, maltooligosaccharide, and amylose.

Antimicrobial alkylamine-modified sugars were prepared. Microwave-assisted click reaction efficiently achieved poly-functionalization of oligo- and polysaccharides. The sugars exhibited a unique relationship of their bacterial membrane permeabilization and antimicrobial activity with the number of functional groups and the structure of the molecular scaffold. It shows that the assembly of the functional groups is necessary for being antimicrobial. The amylose derivatives also exhibited synergy to minimize the necessary dose of conventional antibiotics and increase their antimicrobial potency.

Synthesis and characterization of acetylated amylose and development of inclusion complexes with rifampicin.

Amylose (AM) tends to form single helical inclusion complexes with suitable agents. These complexes are considered promising biomaterial carrier since the guest molecules can be released later, leading to many applications, especially in the pharmaceutical industry. Rifampicin (RIF) has long been recognized as an active drug against Mycobacterium tuberculosis, however, the administration of RIF in high dosages can originate unwanted side-effects. Due to the fact that the use of native amylose (AM) in the formation of complexes is limited by their low water solubility, it was acetylated with a medium degree of substitution (DS), allowing solubilizing (0.5gL-1) acetylated amylose (AMA) in water at neutral pH, in opposition to that observed with native amylose (trace solubility). The resulting acetylated amylose was characterized by means of Fourier Transform Infrared (FT-IR) spectroscopy and Scanning Electron Microscopy (SEM). FT-IR results indicated that the acetylation of anhydroglucose units of amylose corresponds to a low DS, whereas SEM results suggested that the smooth surfaces of amylose granules were changed into rougher surfaces after acetylation. Ultraviolet absorption spectroscopy (UV-vis) analysis confirmed the formation and allowed the quantification of both native (AM-RIF) and acetylated (AMA-RIF) amylose inclusion complexes. Their characterization in solution was performed by dynamic light scattering (DLS) and zeta potential (ZP) measurements. The average size of inclusion complexes as determined by DLS, ranged between 70 and 100nm. Besides, ZP analysis showed that both complexes are more stable in the presence of RIF. This study may lead to the development of an effective method for the preparation of amylose inclusion complexes, which is beneficial to their further application in drug delivery systems.

Synthesis and characterization of amylose grafted poly(acrylic acid) and its application in ammonia adsorption.

Amylose grafted poly(acrylic acid) (Am-g-PAA) was synthesized by graft copolymerization of amylose with acrylic acid. The structure of Am-g-PAA was confirmed by (1)H NMR and FT-IR spectra. The morphology, crystallinity and thermal properties of amylose and Am-g-PAA were investigated by SEM, XRD and TGA, respectively. The highest degree of substitution (DS) of carboxyl group was 1.96 which was obtained after reacted for 1h at 60°C. Acrylic acid to anhydroglucose mole ratio for DS was 19.81. It was found that a large number of carboxyl groups were grafted on the backbone of amylose. It was also found that ammonia adsorption capacity of amylose increased by grafting poly(acrylic acid) on the backbone of amylose.

Synthesis and chiral recognition of amylose derivatives bearing regioselective phenylcarbamate substituents at 2,6- and 3-positions for high-performance liquid chromatography.

Eighteen novel amylose derivatives bearing different phenylcarbamate substituents at 2,6- and 3-positions of a glucose ring were synthesized through the regioselective protection at 2- and 6-positions using a bulky trialkylsilyl chloride. Their chiral recognition abilities were then evaluated as the chiral stationary phases (CSPs) for high-performance liquid chromatography (HPLC) after coating them on the surface of macroporous silica gel. The chiral recognition abilities of these CSPs intricately depended on the nature, position and number of the substituents on the phenyl moieties. The introduction of substituents at meta-position of aromatic moieties at 2- and 6-positions of glucose unit was more attractive than other positions to improve the chiral recognition ability of these amylose derivatives. Each CSP seems to possess its own characteristic resolving power, and those based on amylose 3-(3,5-dichlorophenylcarbamate) showed comparatively better chiral recognition than others. For some racemates, the amylose derivatives with different phenylcarbamate substituents at 2,6- and 3-positions exhibited higher enantioselectivity than the amylose tris(3,5-dimethylphenylcarbamate), which is commercially available as Chiralpak AD, one of the most powerful CSPs. The structures of the obtained amylose derivatives were also investigated by circular dichroism spectroscopy.

Synthesis of Amylose-b-P2 VP Block Copolymers.

A new class of rod-coil block copolymers is synthesized by chemoenzymatic polymerization. In the first step, maltoheptaose, which acts as a primer for the synthesis of amylose, is attached to poly(2-vinyl pyridine) (P2 VP). The enzymatic polymerization of maltoheptaose is carried out by phosphorylase to obtain amylose-b-P2 VP block copolymers. The block copolymer is characterized by Fourier transform infrared spectroscopy, nuclear magnetic resonance, gel permeation chromatography, and wide-angle X-ray scattering techniques. The designed molecules combine the inclusion complexation ability of amylose with the supramolecular complexation ability of P2 VP and therefore this kind of rod-coil block copolymers can be used to generate well-organized novel self-assembled structures.

Thermal expansion behavior of A- and B-type amylose crystals in the low-temperature region.

The thermal expansion behaviors of A-type and B-type amylose crystals, which were prepared by recrystallization of short amylose chains synthesized by phosphorylase, were investigated using synchrotron X-ray powder diffraction between 100 and 300K. For both types of crystals, the room-temperature phase (RT phase), which is the usually observed phase, transitioned to a low-temperature phase (LT phase), on cooling. The phase transitions took place reversibly with rapid changes in the unit-cell parameters around 200-270K. The differences between the RT and LT phase were investigated using solid-state (13)C NMR spectroscopy, which revealed there were changes in molecular chain conformations. These results suggest that the phase transition of water molecules on the crystalline surfaces affects the thermal behavior and structure of polysaccharide crystals.

Facile synthesis and structural characterization of amylose-Fatty Acid inclusion complexes.

Amylose-fatty acid inclusion complexes can be easily prepared by simple mixing in hot aqueous solutions. Above a critical chain length (C6) of the fatty acid insoluble complexes between amylose and each fatty acid (C8, C10, C12, C14, C16) were precipitated from the solution, and characterized by FT-IR, XRD, DSC, and SEC. The presence of the characteristic (CO) FT-IR absorption peak at 1 710 cm(-1) confirmed the inclusion of the fatty acids inside the amylose helix. XRD showed the same characteristic features of the V amylose single helical structure. Both SEC and DSC revealed that longer fatty acids can form inclusion complexes with amylose fractions having higher degree of polymerization, leading to greater yields, and higher thermal stability (higher melting temperature and enthalpy) of the amylose-complexes.

Enzymatic synthesis of amylose brushes revisited: details from X-ray photoelectron spectroscopy and spectroscopic ellipsometry.

The successful synthesis of amylose brushes via enzymatic "grafting from" polymerization and the detailed characterization of all synthetic steps by X-ray photoelectron spectroscopy (XPS) and spectroscopic ellipsometry measurements are reported. Au and Si surfaces are amino-functionalized with self-assembled monolayers (SAMs) of cystamine and 3-aminopropyldimethyethoxysilane (APDMES), respectively. Maltoheptaose is covalently attached to the amino-functionalized Au and Si surfaces via reductive amination. Amylose brushes are grown from maltoheptaose modified Au and Si surfaces with enzymatic polymerization using potato phosphorylase and Rabbit Muscle phosphorylase, as evidenced by spectroscopic ellipsometry and XPS measurements.

Synthesis, characterization, and comparative analysis of amylose-guest complexes prepared by microwave irradiation.

The preparation and characterization of amylose-small molecule complexes is a heavily researched area. There are few reports, however, that compare complexation efficiencies across a matrix of different amylose hosts and guests. We present herein a detailed account of using microwave irradiation to prepare amylose-small molecule complexes in water. Microwave heating reduced the time required to prepare these amylose complexes from hours to minutes. We characterized not only the quantity of complex for each amylose-guest pairing but also the loading of small molecule guest in that complex. Amylose-1-naphthol complexes were found to have the highest loading density compared with other hydrophobic guests studied; in the case of 1-naphthol, there was a linear dependence of guest loading on amylose molecular weight. In addition, complexes featuring 1-naphthol were the most ordered as judged by powder X-ray diffraction (XRD) and differential scanning calorimetry. Further, powder XRD analysis of the microwave-prepared complexes revealed that many contained mixtures of V-form (single helix) and B-form (double helical) amylose. Lastly, untreated Hylon VII complexed the widest variety of small molecules with the overall greatest efficiency.

Synthesis of amylose-polystyrene inclusion complexes by a facile preparation route.

The formation of amylose-polystyrene inclusion complexes via a novel two-step approach is described. In the first-step, styrene was inserted inside the amylose helical cavity, followed by free radical polymerization in the second step. The inclusion complexes were characterized by attenuated total reflection fourier transform infrared spectroscopy (ATR-FTIR), ultraviolet spectroscopy (UV), X-ray diffraction (XRD), atomic force microscopy (AFM), and differential scanning calorimetry (DSC). The formation of polystyrene was confirmed by gel permeation chromatography (GPC). The molecular weight of polystyrene can be varied by using amylose bearing different molar masses. The approach described here is general and could be used to synthesize other host-polymer inclusion complexes for which long chains of polymeric guests are difficult to insert into the host cavity.

SAXS conformational tracking of amylose synthesized by amylosucrases.

Amylose, a linear polymer of α(1,4)-linked glucosyl units and a major constituent of starch granules, can also be enzymatically synthesized in vitro from sucrose by bacterial amylosucrases. Depending on the initial sucrose concentration and the enzyme used, amylose oligomers (or polymers) are formed and self-associate during synthesis into various semicrystalline morphologies. This work describes for the first time a synchrotron SAXS study of the structure in solution of two amylosucrases, namely, NpAS and the thermostable DgAS, under conditions of polymer synthesis and, simultaneously, the amylose conformation. The structure in solution of both amylosucrases during the reaction was shown to be similar to the known crystallographic structures. The conformation of amylose produced at an early stage consists of a mixture of wormlike chains and double helical cylindrical structures. In the case of NpAS, in a second stage, individual double helices pack into clusters before crystallizing and precipitating. Amylose produced by DgAS never self-associates in such clusters due to the higher temperature used for amylose synthesis. All the dimensions determined for wormlike chains and cylindrical conformations at different times of NpAS synthesis are in very good agreement with structural features usually observed on gels of amylose extracted from starch. This provides new insights in understanding the mechanisms of amylose gelation.

Preparation and characterization of new and improved soluble-starches, -amylose, and -amylopectin by reaction with benzaldehyde/zinc chloride.

Seven different starches from potato, rice, maize, waxymaize, amylomaize-VII, shoti, and tapioca, and potato amylose and potato amylopectin have been reacted with benzaldehyde, catalyzed by ZnCl(2), to give new water-soluble starches and water soluble-amylose and soluble-amylopectin. In contrast to the native starches, aqueous solutions of the modified starches could not be precipitated with 2-, 3-, or 4-volumes of ethanol. ß-Amylase gave no reaction with the modified starches, in contrast to the native starches, indicating that the modification occurred exclusively at the nonreducing-ends, giving 4,6-benzylidene-D-glucopyranose at the nonreducing-ends. Reactions of α-amylase with native and modified potato and rice starches gave a decrease in the triiodide blue color and an increase in the reducing-value that were similar for the native- and modified-starches, indicating the modified starches had not been significantly altered by the modification. The benzaldehyde-modified starches and benzaldehyde-modified potato amylose and potato amylopectin components, therefore, have a starch structure very much like their native counterparts, in contrast to the Lintner, Small, and the alcohol/acid-hydrolyzed soluble-starches that have undergone acid hydrolysis. The benzaldehyde-modified starches and starch components have significantly higher water solubility than their native counterparts even though the structures of the modified starches had only been slightly altered from the structures of their native counterparts. They all gave crystal-clear solutions that did not retrograde.

Cationic amylose-encapsulated bovine hemoglobin as a nanosized oxygen carrier.

Nanosized hemoglobin-based oxygen carriers are one of the most promising blood substitutes. In the present study, a comprehensive strategy for the preparation of nanosized cationic amylose-encapsulated hemoglobins (NCAHbs) was developed. First, cationic amylase (CA) was synthesized from amylose and quaternary ammonium salt by an etherification reaction. The structure of CA was characterized using Fourier transform infrared spectrophotometry (FTIR) and proton nuclear magnetic resonance spectrophotometry ((1)H NMR). The degree of substitution and the zeta potential were also measured. Then, the NCAHbs were prepared by electrostatic adhesion, reverse micelles and cross-linking. The UV-visible spectrophotometer was used to measure the entrapment efficiency (EE%) and drug loading efficiency (DL%) of the NCAHbs. Transmission electron microscopy and Malvern Nano-zs 90 analyzer were used to observe the size distribution and morphology of particles. Chemical structure was determined from the FTIR spectrum. A Hemox analyzer was used to measure the P(50) and Hill coefficients. A lethal hemorrhagic shock model in rats was used to evaluate the therapeutic effect of the NCAHbs. The results showed that the combined methods improved the size, stability, EE%, DL%, and oxygen-carrying capacity of the NCAHbs. The average diameter of the NCAHbs was 92.53 ± 3.64 nm, with a narrow polydispersity index of 0.027. The EE% was 80.05% ± 1.56% and DL% was 61.55% ± 1.41%. The P(50) and Hill coefficient were equal to 28.96 ± 1.33 mmHg and 2.55 ± 0.22, respectively. The size of NCAHbs remained below 200 nm for six days in PBS solution. The NCAHbs could effectively prevent lung injury from progressing to lethal hemorrhagic shock because they acted as both a volume expander and an oxygen carrier.

[Synthesis of colon-specific prodrug of indomethacin and its inhibitory effect on liver metastasis from colon cancer].

OBJECTIVE: To develop a colon-specific prodrug of Indomethacin microbially triggered, carry out in vitro/in vivo evaluation of drug release, and appraise its inhibitory effect on liver metastasis from colon cancer. METHODS: Indomethacin prodrugs were synthesized and characterized by FTIR and NMR, and dissolution test simulating gastrointestinal tract was employed to screen the colon-specific prodrug. Then, the pharmacokinetic profile of portal vein and peripheral blood in Sprague-Dawley rats was studied. Lastly, the inhibitory effect on liver metastasis from colon cancer in nude mice was observed. RESULTS: The chemical structure characterized by FTIR and NMR demonstrated that six kinds of indomethacin-block-amylose with different drug loading (IDM-AM-1-6) were synthesized, among which IDM-AM-3 was degraded 1.3%, 9.3% and 95.3%, respectively, in simulated gastric fluid for 4 h, small intestine for 6 h, and colon for 36 h. The pharmacokinetic test of IDM-AM-3 showed that absorption was delayed significantly (P < 0.01), peak time [(11.35 + or - 2.45) h], elimination half-life [(16.74 + or - 4.04) h] and mean residence time [(22.27 + or - 0.52) h] were significantly prolonged (P < 0.01), as well as peak serum concentrations [(9.69 + or - 2.40) mg/L] and AUC(0-t) [(236.7 + or - 13.1) mg x L(-1) x h] were decreased markedly (P < 0.01) as compared with those of IDM regarding to portal vein. Additionally, its AUC(0-t) in peripheral blood was remarkably lower than that in Portal vein (P < 0.01). The tumor suppression observation showed that it could remarkably reduce the number of liver metastases in contrast to IDM (P < 0.05). CONCLUSION: Colon-specific IDM-AM-3 possesses advantage of sustained release in portal vein providing some experimental basis for colon-specific delivery system applied to sustained release in the portal vein.