Characterization of enzymatically synthesized amylopectin analogs via asymmetrical flow field flow fractionation

Asymmetrical flow field flow fractionation (AF4), when coupled with multi-angle laser light scattering (MALLS), is a very powerful technique for determination of the macromolecular structure of high molar mass (branched) polysaccharides. AF4 is a size fractionation technique just as size exclusion chromatography (SEC), nevertheless can overcome some crucial problems found in SEC analysis especially in starch like structures. This paper describes a detailed investigation of the macromolecular structure of two groups of well-defined synthetic amylopectin analogs – synthesized via an in vitro enzyme-catalyzed reaction using the enzymes phosphorylase b from rabbit muscle and Deinococcus geothermalis glycogen branching enzyme (Dg GBE). Size, molar mass distributions and structural data were studied by AF4 coupled with online quasi-elastic light scattering (QELS) and multi-angle light scattering (MALLS).     

Molar mass determination of poly(octadecene-alt-maleic anhydride) copolymers by size exclusion chromatography and dilute solution viscometry

The number average molar mass M_n of poly(octadecene-alt-maleic anhydride) (PODMA) copolymers calculated from data obtained by size exclusion chromatography (SEC) using a polystyrene (PS) calibration was found to be inaccurate. The use of SEC combined with dilute solution viscometry enabled a method to be developed using an iterative approach, which does not require knowledge of the Mark-Houwink constants for PODMA samples. A new calibration curve was constructed as a plot of molar mass M_u for PODMA. True number-average molar masses M_n (true) calculated using the new calibration are approximately twice the apparent molar mass M_n (app) based on a PS calibration for higher molar mass samples (>10?000 g mol⁻¹).     

Extension of the measuring range in size exclusion chromatography to higher molar mass by use of a light scattering detector

A new method for the extension of the measuring range of size exclusion chromatography coupled with light scattering (SEC/LALLS) to higher molar mass is presented. This method uses the fact that the light scattering signal can often be detected already at considerable smaller values of the elution volume (i. e. higher values of molar mass) than the concentration signal; furthermore, it takes into account the peak broadening effect in SEC. This procedure is tested for the case of a logarithmic normal distribution and a most probable distribution of molar mass. In the case of polypropylene with a logarithmic normal distribution (M?_w = 420 000, M?_n = 99 000), the range of molar mass covered by SEC/LALLS can be extended by a factor of about 10; with a most probable distribution (M?_n = 100 000), the attainable extension in molar mass assumes a factor of about 2.     

Characterization of branched polymers by size exclusion chromatography coupled with multiangle light scattering detector. I. Size exclusion chromatography elution behavior of branched polymers

The elution behavior of branched macromolecules during their separation by size exclusion chromatography (SEC) was studied. The elution behavior of branched polymers was investigated using samples of randomly branched polystyrene and star branched poly(benzyl methacrylate) of different levels of branching by means of a SEC chromatograph coupled with a multiangle light scattering detector. Abnormal SEC elution behavior was found to be typical for highly branched polymers. After a normal elution at small elution volumes the molar mass and root mean square radius of the eluting molecules increased with increasing elution volume. Several SEC experiments were carried out to find explanation for this effect and SEC separation was compared with the separation by thermal field flow fractionation. © 2001 John Wiley & Sons, Inc. J Appl Polym Sci 81: 1588–1594, 2001     

Aliphatic tertiary amine mediated synthesis of highly branched polylactide copolymers

In the Sn(Oct)₂ catalyzed bulk copolymerization of l-lactide (LLA) and a large amount of branching comonomer 2,2-bis(hydroxymethyl)butyric acid (BHB) (LLA/BHB/Sn(Oct)₂ ratio is 5:1:0.05, 135 °C), low molecular weight of copolymers with inhomogeneous structure were generated. MALDI-TOF mass spectrum demonstrated that the lower molar mass fraction was dominated with the BHB homo-polycondensation products; whereas the higher molar mass fraction was mainly composed of linear chains of poly(l-lactide) (PLLA) bearing one BHB unit. The addition of aliphatic tertiary amine to this polymerization system could effectively enhance the molecular weight of the obtained copolymers, whereas, aromatic tertiary amines and aliphatic primary and secondary ones were not so effective. The obtained PLLA copolymers were characterized by GPC, ¹H NMR, ¹³C NMR and MALDI-TOF MS, which verified that they were composed of LLA and BHB units and had the highly branched structure. The degree of branching was about 0.23–0.30. Thermal analyses by TGA and DSC demonstrated that the resulting highly branched PLLA copolymers were amorphous. Compared with linear PLLA, the highly branched PLLA copolymers were thermally more stable and had lower T_g. The mechanism of aliphatic tertiary amine mediated synthesis of medium molecular weight of highly branched PLLA copolymers was proposed.     

Synthesis and Characterisation of Branched Poly(ethylene terephthalate)

Branched poly(ethylene terephthalate)s (PET) were synthesised with a variety of molar masses and with a large range of degree of branching by introduction of mono-, tri-(glycerol) and tetra-functional (pentaerythritol) comonomers to dimethyl terephthalate and ethylene glycol. The monofunctional alcohols, dodecanol and benzyl alcohol, were used as terminating agents to minimise gelation. The effect of various reaction parameters, such as percentage glycerol or pentaerythritol and polymerisation time, on limiting viscosity number [η] and weight average molar mass (M_w) were investigated. The thermal behaviour of branched PET was studied by differential scanning calorimetry; all samples showed a characteristic double endothermic melting peak and the glass transition temperature was not observed. Some branched PETs were subjected to solid-state polymerisation to increase the molar mass of previously prepared branched polymers. The solid-state polymerisation technique showed that the process not only promoted the molar mass but, more importantly, it increased the crystallinity of the polymer. Overall, the solid-state reaction rate was governed by initial molar mass, crystallinity, reaction temperature and time. © of SCI.     

Reactive extrusion of acrylic acid grafted polypropylene with hexadecylamine

To study the possibility of the production of branched polypropylene (PP) by a reactive extrusion (REX) route, side chains were introduced on the backbone of a polypropylene material by reacting hexadecylamine with acrylic acid grafted PP. Experiments were carried out both in solution and in the melt, and the products were analyzed by FTIR, elemental analysis, dynamic mechanical, and rheological techniques. Analysis of the FTIR spectra of the samples produced in the solution reactions, at an equal molar ratio of [ –NH₂]/[–COOH] without catalyst addition and without removal of the by-product, revealed that the formation of imide was increased with increasing the reaction time up to 10 h, while a further increase in reaction time resulted in a reversal of the reaction. In the REX experiments, FTIR analysis showed that the imide formation increased with the [–NH₂]/[–COOH] molar ratio. At a molar ratio of one, more imide was present in the REX product than the in-solution one. Elemental analysis suggested that the nitrogen content in the products initially increased with [–NH₂]/[–COOH] molar ratio and then reached an almost constant value at molar ratio values of about unity. The glass transition temperature (T_g) was measured by dynamic mechanical analysis (DMA), and it was found that the attachment of the alkyl chains caused a reduction in T_g of the products. Finally, rheological measurements showed that the shear viscosity of the products increased with the amine/carboxyl molar ratio at low shear rates and that their moduli were enhanced as a result of the attachment of the alkyl side chains.     

Purification and Characterization of Microbial Hyaluronic Acid by Solvent Precipitation and Size-Exclusion Chromatography

Abstract

Samples of hyaluronic acid (HA) produced by submerged fermentation using a synthetic culture medium were recovered, purified, fractioned, and characterized using solvent precipitation and size-exclusion chromatography (SEC). The samples showed a wide molar mass distribution in the range 10³–10⁷ Da, most of which had an average molar mass between 10⁴ and 10⁵ Da after purification by sequential precipitations. Fractions of HA with molar mass above 10⁵ Da were purified by SEC in a semi-preparative scale with nearly no protein contamination. Characterization and fractionation of the HA was carried out by SEC in a analytical and semi-preparative scale using Shodex OHPak SB806M HQ and Superose 6 columns, respectively.     

Chemistry of Secondary Organic Aerosol Formation from OH Radical-Initiated Reactions of Linear,Branched, and Cyclic Alkanes in the Presence of NO x

The effects of molecular structure on the products and mechanisms of SOA formation from OH radical-initiated reactions of linear, branched, and cyclic alkanes in the presence of NO _x were investigated in a series of environmental chamber experiments. SOA mass spectra were obtained in real time and off line using a thermal desorption particle beam mass spectrometer and used to identify reaction products. Real-time mass spectra were used to classify products according to their temporal behavior, and off-line temperature-programmed thermal desorption analysis of collected SOA was used to separate products by volatility prior to mass spectral analysis and to gain information on compound vapor pressures. A reaction mechanism that includes gas- and particle-phase reactions was developed that explains the formation of SOA products and is consistent with the various lines of mass spectral information. Results indicate that the SOA products formed from the reactions of linear, branched, and cyclic alkanes are similar, but differ in a few important ways. Proposed first-generation SOA products include alkyl nitrates, 1,4-hydroxynitrates, 1,4-hydroxycarbonyls, and dihydroxycarbonyls. The 1,4-hydroxycarbonyls and dihydroxycarbonyls rapidly isomerize in the particle phase to cyclic hemiacetals that then dehydrate to volatile dihydrofurans. This conversion process is catalyzed by HNO ₃ formed in the chamber and is slowed by the presence of NH ₃ . Volatile products can react further with OH radicals, forming multi-generation products containing various combinations of the same functional groups present in first-generation products. For linear and branched alkanes, the products are acyclic or monocyclic, whereas for cyclic alkanes they are acyclic, monocyclic, or bicyclic. Some of the products, especially those formed from ring-opening reactions of cyclic alkanes appear to be low volatility oligomers. The implications of the results for the formation of atmospheric SOA are discussed.     

Molar mass distribution by size exclusion chromatography: Comparison of multi-angle light scattering and universal calibration

Series of polymers of various molar mass, chemical composition, and molecular architecture was analyzed by size exclusion chromatography (SEC) coupled with a multi-angle light scattering (MALS) photometer and an online viscometer. The molar mass averages were determined from the signal of MALS or calculated from the intrinsic viscosity and universal calibration. The comparison of the obtained results showed significant differences between the two methods. The MALS detection was shown to be more accurate for the determination of the weight-average molar mass and less vulnerable to the spreading of polymer peak by band broadening. The universal calibration can yield more accurate estimation of the number-average molar mass of branched polymers. It is also significantly more accurate for the characterization of fluorescent polymers than MALS with a regular laser of 660 nm. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2019 , 136, 47561.     

Topological separation of linear and star-shaped polystyrenes by off-line 2D chromatography. Stars having high molar mass arms and quantification of the star fraction

Off-line 2D separations on mixtures of linear and star-shaped polymers were performed using temperature gradient interaction chromatography (TGIC) as a first and SEC as a second dimension. The experiments resulted in clear separations of the linear and star shaped-structures for arm molar masses up to 42,000 g/mol. The resolution is nearly independent of the molar masses of the arms and depends only on the number of star arms. From the 2D chromatograms it is possible to determine the molar mass of the first branched structure, i.e. the three arm star. The evaluation of the relative peak volume allows a reliable estimation of the amount of branched structures in the complex mixture.     

Molar mass of poly (ethylene terephthalate) (PET) during ultimate uniaxial drawing

The changes of the average molar mass M_w, M_n, M_z, and molar mass distributions during multistep uniaxial drawing of poly(ethylene terephthalate) (PET) to achieve ultimate mechanical properties have been studied in detail by means of size exclusion chromatography (SEC) with triple detection: concentration, viscosimetry, and light scattering, using HFIP as solvent. An increase in molar mass of PET due to post‐polycondensation and/or transesterfication during drawing at a high temperature of 160 to 230°C was found. Moreover, drawing leads to crystallization and large orientation in the amorphous phase, which results in lower molecular mobility and prevents a further growth in chain length. Crazing under extreme drawing conditions occurs and affects a decrease in molar mass.     

Hydroformylation of unsaturated fatty esters

Unsaturated fatty esters and vegetable oils were hydroformylated with H₂ and CO (3500–4600 psi) and Co₂(CO)₈ to give fatty aldehydes at 100–110 C and fatty alcohols at 175–190 C. Yields of distillable C₁₉ oxo products varied from 42% to 84%. Distilled products contained from 50% to 90% branched isomers and from 4% to 16% linear isomers. The proportion of linear isomers increased at higher reaction temperatures and in the presence of tributylphosphine-cobalt carbonyl complex. Linear and branched hydroxy products were separated by silicic acid column, thin-layer, and gas-liquid chromatography. The linear hydroxy product (from oleate and linoleate) was identified as methyl 19-hydroxynonadecanoate by nuclear magnetic resonance and mass spectrometry. Isomeric branched products were analyzed by mass spectrometry as the diester derivatives. They were identified as a mixture of 5- to 13-carbomethoxy methyl octadecanoate. Presented at the AOCS-AACC Joint Meeting, Washington, D.C., March 1968. No. Utiliz. Res. Dev. Div., ARS, USDA. Bureau of Mines, U.S. Dept. of Interior.     

Molar mass distribution of polybutadiene synthesized with nickel-based Ziegler-Natta catalysts,II

Cis-1,4-polybutadiene was produced by polymerization of 1,3-butadiene using a catalyst system containing Ni(OOC₈H₁₅)₂ and Al(C₂H₅)₃ and the Lewis acids BF₃ · O(C₂H₅)₂ or TiCl₄. The molar mass distributions (MMD) of the polybutadienes are investigated by means of size exclusion chromatography (SEC). The MMD's were fitted by a sum of Schulz-Flory-Functions (SFF). Taking into account that one kind of active species gives one SFF defined by molar mass averages, one comparable active species in both systems and one different were found. This way it was tried to find a relationship between the grown up of the active species depending on components and reactions conditions.     

C18‐unsaturated branched‐chain fatty acid isomers: Characterization and physical properties

Iso‐oleic acid is a mixture of C₁₈‐unsaturated branched‐chain fatty acid isomers with a methyl group on various positions of the alkyl chain, which is the product of the skeletal isomerization reaction of oleic acid and is the intermediate used to make isostearic acid (C₁₈‐saturated branched‐chain fatty acid isomers). Methyl iso‐oleate, a mixture of C₁₈‐unsaturated branched‐chain fatty acid methyl ester isomers, is obtained via acid catalyzed esterification of iso‐oleic acid with methanol. The branched‐chain materials are liquid at room temperature and their “oiliness” property makes them an attractive candidate for the lubricant industry. In this paper, we report characterization of these branched‐chain materials using comprehensive two‐dimensional GC with time‐of‐flight mass spectrometry (GC × GC/TOF‐MS) and their physical and lubricity properties using tribology measurements.     

Size-exclusion chromatography with two-angle laser light-scattering (SEC-TALLS) of high molecular weight and branched polymers

The accuracy and precision of results obtained from light-scattering detection at two angles (TALLS) for size-exclusion chromatography (SEC) are examined for linear narrow molecular weight distribution polystyrenes between 1,290,000 and 20,000,000 MW and for branched polyesters. The ratio of light-scattering intensities at 15° and 90° is used to calculate weight-average molecular weight, M?_w, and an average root-mean-square radius, r?_gu, equivalent to the z-average radius. A shape for the polymer molecule is assumed and an analytical relationship for the particle-scattering function is required. It is shown that analysis of the data using the particle-scattering function for a random coil is valid for both high molecular weight, linear polystyrenes and long-chain branched polyesters. The radius, r?_gu, is determined with high precision by using the ratio of light-scattering signals, which is insensitive to errors in sample concentration and changes in the eluent flow rate. The correct average radius for the whole polymer is obtained despite using low-efficiency, large-particle diameter SEC columns; however, axial dispersion significantly affects molecular weights and radii calculated at each retention volume that can limit the utility of plots used to deduce polymer conformation. © 1995 John Wiley & Sons, Inc.     

Characterization of poly (N‐vinyl formamide) by size exclusion chromatography–multiangle light scattering and asymmetric‐flow field‐flow fractionation–multiangle light scattering

The molar mass and the radius of gyration of three poly N‐vinyl formamide (polyNVF) synthesized in aqueous solution polymerization were characterized using two different fractionation techniques: size exclusion chromatography (SEC) and asymmetric‐flow field‐flow fractionation (AF4) coupled with a multiangle light scattering (MALS) and a refractive index (RI) detector. For the sake of comparison, the polymers were also characterized by MALS using the Zimm plot approach (no fractionation). The dn dc^?1 of the poly (N‐vinyl formamide) was measured (0.1564 mL g^?1) and it was found to be insensitive to the molar mass (in the range 150–450 kDa) and also to the eluents used (DDI water or mixed eluent DDI water/acetonitrile (80 : 20) at pH = 5.5). Interestingly, the concentrations of the samples injected in the SEC and AF4 should be different because concentrations in the range of 20–40 mg mL^?1 used for the AF4 caused overloading and anomalous elution in the SEC and hence misleading molar masses. At adequate concentrations in each fractionation equipment, the molar masses were in reasonable good agreement although AF4/MALS provided larger values than the other two techniques likely because samples were not filtered before injection. © 2015 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2015 , 132, 42434.     

Polycardanol's potential to deasphalt crude oil: Influence of polymer conversion degree,molar mass,and structure

Cardanol is a natural material that acts to stabilize asphaltenes in crude oil. However, its derivative, polycardanol, obtained via cationic polymerization, has divergent behaviour, with reports of its acting both as asphaltenes stabilizer and flocculant. Recently, it was demonstrated that the reaction conditions influence the conversion rates, structures, and molar masses of the products from synthesizing polycardanol initiated with BF₃ · O(C₂H₅)₂. Seeking to elucidate the influence of these variables on the phase behaviour of asphaltenes, in this work six products, previously synthesized and characterized, had their performance evaluated by asphaltenes precipitation onset, using n-heptane titration and monitoring by near-infrared spectroscopy, using asphaltenes model systems (C5I and C7I) at 1.00 wt./vol.% in toluene. All the products had the ability to flocculate asphaltenes and have potential for deasphalting process. Flocculation efficiency increased with rising molar mass, higher reaction conversion degree, and presence of lateral hydrocarbon chains. The most efficient structures were not affected by the presence of unreacted cardanol. This indicated that the reaction does not require a purification step. Aged cardanol was also able to produce flocculant polymer, indicating that the distillation of cardanol is not required. The cost to obtain a product without needing reagent distillation becomes lower.     

Lithium naphthalenides in non-polar or in low-polarity media: some insights regarding their use as initiators in anionic polymerizations

The synthesis of bidirectional anionic initiators by the reaction between metallic lithium (Li) and naphthalene (Naph), under mild conditions, in non-polar (benzene) or low-polarity media (benzene/THF mixtures) is reported. The efficiency of these initiators to provide macromolecules with well-defined structures was demonstrated. Model linear homopolymers from styrene (S) or hexamethyl(ciclotrisiloxane) (D₃) monomers were synthesized using classical anionic polymerization (high-vacuum techniques). The model polymers obtained were analyzed using the conventional analytical techniques, and showed narrow molar mass distributions, a broad range of molar masses (from 3000 to 1,000,000 g/mol) and polydispersity indexes (M _w/M _n) lower than 1.1. High molar mass polymers were obtained using pure benzene as solvent, whereas lower molar masses were obtained in benzene/THF mixtures in which the concentration of THF was lower than 10 % v/v. The ratio [Li]/[Naph] and the nature of the reaction medium are the experimental parameters to be controlled to obtain the desired lithium naphthalenides.     

Full copolymer characterization by SEC-NMR combined with SEC-MALDI

Measurements on a series of random copolymers with units of methylmethacrylate, butylacrylate, styrene and maleic-anhydride are performed. A method is used which consists in fractionating the copolymer by size exclusion chromatography (SEC), collecting 30-40 fractions and then recording both the NMR and MALDI spectra of the fractions. In a successive step, bivariate distribution of chain sizes and composition maps are derived from knowledge of the molar mass, weight and composition of the copolymer fractions. The compositional distribution is obtained by summation of the sections of the bivariate distribution which belong to a narrow compositional range.A model for SEC fractionation of copolymers is developed. It allows to predict the composition and D(x) of the SEC fraction (D(x) is the ratio between the number-average and the weight-average molar mass, x is the fraction number).Offline SEC-MALDI measurements are performed, in order to measure experimentally D(x) for the random copolymer. The agreement between theoretical and experimental values for D(x) and for the composition of the SEC fraction (as derived from SEC-NMR measurements) is good. The predictions of the model were also compared with literature data for SEC fractionation of a copolyester with units of butileneadipate and butilenesebacate. The agreement between theoretical and experimental values is more than satisfactory.