Beate Pfannemüller

Phosphorolytic syntheses with di-, oligo- and multi- functional primers

Abstract Studies on preparation of linear, star-, and comb-shaped polymers carrying amylose chains of uniform length grown by synthesis with potato phosphorylase are reported. Binding of malto-oligosaccharide primers (A) to different supports (B) was performed by the reaction of aldonolactone or N -(2-aminoethyl)aldonamide derivatives of malto-oligosaccharides with carriers respectively containing NH 2 or CO 2 H groups. Synthetic products were converted into the tricarbanilate derivatives, and their hydrodynamic behavior was studied by light-scattering and viscosity measurements. The molecular-weight distribution was examined by 10-MPa 1.c. Phosphorolytic syntheses with difunctional primers,. A-B-A [with B = oligo(methylene) or oligo(oxyethylene)], proceed in the same way as with monofunctional primers. Both chain ends are elongated equally to amyloses of uniform length. Oligofunctional primers, (A) x B with x = 3, 4, and 6, lead to a mixture of components differing in the number of long branches, but with the amylose chains all having approximately uniform lengths. Compounds with two branches are preferentially formed. The results are discussed with regard to ( a ) the restrictions of the potato phosphorylase and ( b ) a model in which two primer ends from the oligofunctional substrate molecule are simultaneously bound to the active centers of the phosphorylase dimer. Compared with these densely packed primers, malto-oligomers attached to a linear backbone chain of poly(vinylamine) are attacked more easily. This can be explained by a greater flexibility of the poly(vinylamine) chain.

Amphiphilic properties of synthetic glycolipids based on amide linkages. I. Electron microscopic studies on aqueous gels

Abstract Amphiphiles with one or two amide linkages have been prepared by the reaction ( A ) of D-gluconic acid lactone with aliphatic amines ( C 6 - C 10 ) and (C) of N ′-gluconoyl-ethylenediamine with alkanoic acids ( C 6 )-( C 10 ). Gel formation was found to occur on cooling the aqueous solutions at concentrations as low as 1–2%. Electron microscopy revealed that the gels of type A are composed of highly ordered ropes with right-handed twist, especially well developed with N -octylgluconamide. Type c substances with two amide linkages of opposite direction form gels consisting of smooth ribbons devoid of twisting. N -Methylation of the amide bond (type B and D substances) leads to a considerable increase in solubility. Gels are only formed from samples containing decanoic acid. These gels also consist of right-handed fibrillar ropes, only partially ordered with one N -methylated amide linkage ( B ), regularly aligned side-by-side with one N -methylated and one non-methylated amide bond ( D ). Gel formation and the typical morphology of the gels are discussed as arising mainly from strong intermolecular hydrogen bonds between amide linkages holding the molecules together and the influence of chiral centers of the carbohydrate chain which might be responsible for helical aggregates to be formed.

Influence of chain length of short monodisperse amyloses on the formation of A- and B-type X-ray diffraction patterns

Abstract Amyloses of uniform length were obtained by phosphorlytic synthesis (DP 20–35) and by preparative g.p.c. fractionation by an α-amylolytic digest of amylose on Bio-Gel P-4 (DP 3–20). Crystalline precipitates formed from pure aqueous solution on standing at ambient or lower temperatures gave the A-type X-ray pattern for malto-oligomers of DP 10–12 and the B-type pattern for DP 13 and all longer chains. With these carefully purified samples a mixture of the A- and B-amylose pattern was not observed. Malto-oligomers shorter than DP 10 did not crystallize. The findings support recent studies, indicating that the average chain length of amylopectin and the X-ray type of various starches are closely related. The reason for an influence of chain length on crystalline packing remains to be resolved.

Amphiphilic properties of synthetic glycolipids based on amide linkages. II. Crystal and molecular structure of N-(n-octyl)-D-gluconamide, an amphiphilic molecule in head-to-tail packing mode

Abstract The crystal structure of N-(n-octyl)-D-gluconamide (space group P21, a = 5.252(1), b = 32.426(9), c = 4.805(1) A , β = 94.96(5)° ) was determined by X-ray diffraction methods and refined to R = 0.046. The molecules are V-shaped and occur in an all-trans conformation for both the aliphatic and D-glucose moieties. The crystal structure shows extensive intra- and intermolecular hydrogen bonding and the molecules are packed parallel in sheets, adjacent sheets being arranged in a very unusual head-to-tail fashion. Several homologues of N-(n-octyl)-D-gluconamide were also crystallized. Compounds with an even number of carbon atoms in their alkyl chain crystallize in monoclinic space-group P21, whereas those with an odd number of alkyl carbons occur in triclinic P1. At temperatures between 70°C and 100°C, all compounds undergo a phase transition in the crystalline state as seen in the polarizing microscope. The structure of an octylgluconamide crystal that was heated above the transition temperature and then cooled to room temperature was determined independently and refined to R = 0.071. The structure turned out to be largely identical to the ‘native’ one. Phase transitions were also observed in solution by small angle scattering experiments. These are discussed in relation to the formation of gels which has been reported for these compounds in a previous publication (Pfannemuller and Welte, Chem. Phys. Lipids, 37 (1985) 227–240).

Synthesis of comb-like derivatives of amylose and cellulose having (1→6)-linked d-glucose side-chains

Abstract D -Glucose side-chains were introduced into amylose and cellulose by condensation of tetra- O -acetyl-α- D -glucopyranosyl bromide with 6-trityl-2,3-dicarbanilate derivatives of the polysaccharides in nitromethane- p -dioxane in the presence of silver perchlorate (reaction A ), and with the detritylated derivatives in acetonitrile- p -dioxane in the presence of mercuric cyanide and mercuric bromide (reaction B ). The procedures were equally effective. The distance between the branch points in amylose was in the range 2–4 D -glucose residues, and 5–8 D -glucose residues in cellulose. Whereas considerable degradation of the backbone chain was found with reaction A , no degradation occurred in reaction B under suitable conditions. α-(1→6)-Links were preferably formed in reaction B , and β-(1→6)-links in reaction A .

Comb-like derivatives of amylose having (1-6)-linked malto-oligosaccharide side-chains.

The preparation of acetylated glycosyl bromide derivatives of the higher malto-oligosaccharides was studied by using beta-cyclodextrin or linear malto-oligosaccharides of d.p. 6 and 7. The products were treated with 2,3-di-O-phenylcarbamoyl-6O-tritylamylose in the presence of silver perchlorate (reaction A), and with 2,3-di-O-phenylcarbamoylamylose in the presence of mercuric cyanide and mercuric bromide (reaction B). After removal of the substituents, the branched molecules were characterized by their iodine-binding properties, beta-amylolytic degradation, and their priming activity in phosphorolytic synthesis. The distance between the branch points of the backbone chain was 25-55 and 100-150 units in the products from reactions A and B, respectively. Thus, the frequency of branching was considerably lower than in the comb-like molecules having D-glucose side-chains previously described.

Attachment of (1→6)-linked d-glucose side-chains to amylose and cellulose via 1,2-orthoesters

Abstract Comb-like derivatives of amylose and cellulose having (1→6)-linked d -glucose side-chains were prepared by condensation of 1,2-(tert-butyl orthoacetate) and 1,2-(ethyl orthoacetate) derivatives of 3,4,6-tri-O-acetyl-α- d -glucopyranose with 2,3-di-O-phenylcarbamoyl derivatives of amylose and cellulose in boiling chlorobenzene in the presence of catalytic amounts of 2,6-lutidinium perchlorate. The more-stable ethyl orthoacetate yielded a higher and more reproducible degree-of-branching than the tert-butyl orthoacetate. The distance between the branch points in the products was 2-3 d -glucose residues when 1 mol. of orthoester was used for each hydroxyl group in the polysaccharide. Little degradation of the backbone occurred, as shown by intrinsic viscosities and molecular weights determined by light-scattering measurements. Degradation increased with increasing molar ratio of orthoester. The branches were attached mainly (1→6)-β with a small proportion of (1→6)-α linkages as indicated by n.m.r. and i.r. spectroscopy.

H.p.l.c. of 4-nitrophenyl-α-d-malto-oligosaccharides

Abstract The distribution of 4-nitrophenyl-α- d -malto-oligosaccharides produced by phosphorolytic synthesis was analyzed by an amine-modified and a C 18 -bonded silica gel. With both systems high resolution of components up to d.p.40 can be achieved by gradient elution using acetonitrile-water and water-methanol, respectively. The use of water-acetonitrile in reversed-phase (C 18 ) chromatography allows only a separation up to d.p. 9. The elution behaviour of oligomers in the d.p. range of 1–10 glucosyl residues per molecule eluted by water-methanol is discussed. The observation of rearrangement products of 4-nitrophenyl-α- d -malto-oligomers was successful by normal-phase chromatography.

Ordered arrangements in solutions of amylose-iodine complexes derived from free and terminally fixed amylose chains

Abstract Linear, homodisperse amyloses and branched and cross-linked products are characterized by absorption and c.d. measurements of their iodine complexes in the range 400–780 nm. The long-wave Cotton effect splits into a broad, negative band and a positive band of smaller width. Dichroic absorption has maximum values at d.p. values of about 50 for iodine concentrations of 0.7, 1.0, and 1.4 I − 3 per six glucose residues. A rather stiff, helical rod is suggested for this chain length. Compared with free amyloses of equal length, amylose branches linked densely to glycogen give stronger Cotton effects, thus indicating a significantly ordered arrangement. Amylose segments in soluble, cross-linked gels, when compared with linear amyloses of equal length, show 60% of the iodine-binding capacity, but only 20–25% of the dichroic absorption. This effect is due to imperfect helix-formation. From experiments under various conditions (amylose concentration, rapid and slow addition of iodine), there is strong evidence that the time-dependent increase of c.d. results mainly from intramolecular orientation rather than from intermolecular association. The formation of larger aggregates may be ruled out, under the conditions applied.

Attachment of (1→6)-linked malto-oligosaccharide side-chains to amylose and cellulose via 1,2-orthoesters

Abstract Comb-like molecules having oligomeric side-chains were prepared by condensation of peracetylated 1,2-(ethyl orthoacetate) derivatives of maltose, maltotetraose, and maltohexaose with 2,3-di- O -phenylcarbamoyl derivatives of amylose and cellulose. The distance between the maltose, maltotetraose, and maltohexaose branch-points introduced into amylose was 3–4, 10, and ∼ 17 d -glucose residues, respectively, and only slightly greater for cellulose. Thus, the branch frequency is considerably higher than that obtained previously by using the Bredereck and Helferich reactions. After saponification of the modified polysaccharides, the side chains of d.p. 4 and 6 attached to position 6 in the d -glucose residues of amylose and cellulose were easily extended by enzymic synthesis with potato phosphorylase.