Richard H. Furneaux

Some reactions of levoglucosenone

Abstract The role of the major variables in the pyrolytic production of levoglucosenone from acid-treated cellulose and paper has been established. Reduction, oxidation, acid- and base-catalyzed addition, acetolysis, and rearrangement reactions of this product have been investigated. These reactions proceed with considerable stereo-selectivity to give a variety of novel compounds. Nucleophilic addition to the double bond provides a facile method for the synthesis of 4-thio and other 4-substituted sugar derivatives.

Immucillin H, a powerful transition-state analog inhibitor of purine nucleoside phosphorylase, selectively inhibits human T lymphocytes.

Transition-state theory has led to the design of Immucillin-H (Imm-H), a picomolar inhibitor of purine nucleoside phosphorylase (PNP). In humans, PNP is the only route for degradation of deoxyguanosine, and genetic deficiency of this enzyme leads to profound T cell-mediated immunosuppression. This study reports the biological effects and mechanism of action of Imm-H on malignant T cell lines and on normal activated human peripheral T cells. Imm-H inhibits the growth of malignant T cell leukemia lines with the induction of apoptosis. Imm-H also inhibits activated normal human T cells after antigenic stimulation in vitro. However, Imm-H did not inhibit malignant B cells, colon cancer cell lines, or normal human nonstimulated T cells, demonstrating the selective activity of Imm-H. The effects on leukemia cells were mediated by the cellular phosphorylation of deoxyguanosine and the accumulation of dGTP, an inhibitor of ribonucleotide diphosphate reductase. Cells were protected from the toxic effects of Imm-H when deoxyguanosine was absent or when deoxycytidine was present. Guanosine incorporation into nucleic acids was selectively blocked by Imm-H with no effect on guanine, adenine, adenosine, or deoxycytidine incorporation. Imm-H may have clinical potential for treatment of human T cell leukemia and lymphoma and for other diseases characterized by abnormal activation of T lymphocytes. The design of Imm-H from an enzymatic transition-state analysis exemplifies a powerful approach for developing high-affinity enzyme inhibitors with pharmacologic activity.

Carrageenan from the tetrasporic stage of Gigartina decipiens (Gigartinaceae, Rhodophyta)

The structure of the polysaccharide isolated from tetrasporophytic plants of the New Zealand red alga Gigartina decipiens has been determined by chemical and spectroscopic techniques. It is a linear polymer composed primarily of alternating 3-linked beta-D-galactopyranosyl 2-sulphate and 4-linked alpha-D-galactopyranosyl 2,6-disulphate residues. About 15% of the 3-linked residues have an additional sulphate ester group at the 6-position. Aside from this small extra sulphate substitution, the structure is that of the idealised lambda-carrageenan. Good quality solution-state 13C NMR spectra were recorded and interpreted for this carrageenan and for the carrageenans produced from it by solvolytic desulphation and alkali modification.

Structure and antiviral activity of the galactofucan sulfates extracted from Undaria pinnatifida (Phaeophyta)

The galactofucan sulfate extract (GFS) obtained from the brown seaweed Undariapinnatifida by extraction with dilute acid is a potent inhibitor of the herpes viruses HSV-1, HSV-2 and HCMV, with IC50 values determined in vitro of 1.1, 0.2 and 0.5 μgmL−1, respectively. Fractionation of GFS by anion exchange chromatography gave three fractions which differed in their uronic acid and sulfate contents and in their antiviral activity, as well as in having somewhat reduced molecular weights compared to GFS. The low uronic acid/high sulfate fraction (F2M), obtained in 63% yield, had similar molar proportions of galactopyranosyl and fucopyranosyl residues, little associated protein and was equipotent with GFS (IC50 values of 1.1, 0.1 and 0.5 μgmL−1, respectively). The high uronic acid/low sulfate fraction (F1M), obtained in 18% yield, had a much lower proportion of galactopyranosyl residues and was less active (IC50 values of 4.6, 1.0 and 4.0 μgmL−1, respectively). The minor low uronic acid/high sulfate fraction (F4M) had a significant amount of associated protein and was also less active (IC50 = 3.1, 1.0 and 2.0 μgmL−1, respectively). The structure of the major fraction (F2M) was shown to be complex by glycosyl linkage analysis before and after solvolytic desulfation, with many component sugar residues being identified, although 3-linked fucopyranosyl 2,4-disulfate residues were a prominent feature.

Solid-state NMR studies on the structure of starch granules

Abstract Resonances expected for carbon atoms in the amorphous regions of moistened starch granules are not visible in 13C CP/MAS NMR spectra. These resonances could be observed in spectra obtained by using a single-pulse, Bloch-decay sequence, with magic-angle spinning (SP/MAS). This implies that the amorphous regions in moistened starch granules are mobile and have properties similar to a rubberlike polymer. For moistened wheat starch granules, proton rotating frame relaxation times for resonances assigned to the amylose-lysophospholipid inclusion complexes were shorter than those of the crystalline starch. Relaxation differences were used to generate subspectra of the crystalline starch and the amylose-lysophospholipid inclusion complex. Our results suggest that the inclusion complexes occur in distinct regions of the starch granule. There are thus three distinct components making up the wheat starch granule: (i) highly crystalline regions formed from double-helical starch chains, (ii) solid-like regions formed from lipid inclusion complexes of starch, and (iii) completely amorphous regions, associated with the branching regions of the amylopectin component of starch and possibly the lipid-free amylose. All these components can be observed separately using different solid-state NMR techniques and would be expected to contribute to the physical properties of a wheat starch granules.

1,5-Anhydro-4-deoxy-d-glycero-hex-1-en-3-ulose and other pyrolysis products of cellulose

Abstract Uncatalyzed pyrolysis of cellulose provides a tar containing mainly 1,6-anhydro- d -glucose derivatives and some unsaturated products. The latter include a new enone that has been isolated by preparative column chromatography in 1.4% yield and identified as 1,5-anhydro-4-deoxy- d - glycero -hex-l-en-3-ulose. This compound is also formed by pyrolysis of other carbohydrate polymers. A mechanism for its production from internal units has been deduced from the experimental data. The pyrolysis products of cellulose also contain 3,5-dihydroxy-2-methyl-4 H -pyran-4-one, which appears to be an oxidation product.

Structural analysis of carrageenans from the tetrasporic stages of the red algae, Gigartina lanceata and Gigartina chapmanii (Gigartinaceae, Rhodophyta)

The use of modern analytical techniques has facilitated the identification of the polysaccharides from tetrasporophytic Gigartina lanceata and Gigartina chapmanii. The G. lanceata polysaccharide corresponds, very closely, to the idealised structure of λ-carrageenan ([G2S-D2S,6S]n). That from G. chapmanii contains predominantly ξ-carrageenan ([G2S-D2S]n), but with about one in seven of the 3-linked units also has a pyruvate acetal group at the 4- and 6-positions, one in four of the 4-linked units is also sulfated at the 6-position, and a small but significant number of 4-linked 3,6-anhydrogalactosyl 2-sulfate units are present.

Transition-state analogs as inhibitors of human and malarial hypoxanthine-guanine phosphoribosyltransferases

The proposed transition state for hypoxanthine-guanine phosphoribosyltransferases (HGPRTs) has been used to design and synthesize powerful inhibitors that contain features of the transition state. The iminoribitols (1S)-1-(9-deazahypoxanthin-9-yl)-1,4-dideoxy-1,4-imino-D-ribitol 5-phosphate (immucillinHP) and (1S)-1-(9-deazaguanin-9-yl)-1,4-dideoxy-1,4-imino-D-ribitol 5-phosphate (immucillinGP) are the most powerful inhibitors yet reported for both human and malarial HGPRTs. Equilibrium binding constants are >1,000-fold tighter than the binding of the nucleotide substrate. The NMR spectrum of malaria HGXPRT in the Michaelis complex reveals downfield hydrogen-bonded protons. The chemical shifts move farther downfield with bound inhibitor. The inhibitors are lead compounds for species-specific antibiotics against parasitic protozoa. The high-resolution crystal structure of human HGPRT with immucillinGP is reported in the companion paper.

Near-quantitative production of fatty acid alkyl esters by lipase-catalyzed alcoholysis of fats and oils with adsorption of glycerol by silica gel

Abstract A simple solution to the problem of incomplete reaction in lipase-catalyzed alcoholysis of triacylglycerol oils has been found. When immobilized Mucor meihei lipase was used to catalyze the reaction of tallow with three molar equivalents of butanol, the yield of butyl esters did not exceed 70% (w/w). If silica gel was added to the reaction mixture, it adsorbed the glycerol produced during the reaction and the yield increased to up to 98%. The optimum amount of silica gel was 1.25 weight equivalents to the glycerol produced. Other adsorbents were tested, e.g., cellulose, starch, charcoal, and celite, but none was as effective as silica gel. A variety of fats and oils could be used and all gave yields over 90%. Aliphatic alcohols varying in polarity between methanol and dodecanol also gave yields over 90%, although the highest yields were obtained from the most hydrophobic alcohols. The reaction could be carried out by circulating the reaction mixture through columns of enzyme and silica gel. Dehydration of the reaction mixture, and hence of the enzyme, by the silica gel caused loss of activity during repeated reuse of the enzyme. This problem could be minimized by increasing the water content of the initial reaction mixture.

Third-Generation Immucillins: Syntheses and Bioactivities of Acyclic Immucillin Inhibitors of Human Purine Nucleoside Phosphorylase

ImmH (1) and DADMe-ImmH (2) are potent inhibitors of human purine nucleoside phoshorylase (PNP), developed by us and currently in clinical trials for the treatment of a variety of T-cell related diseases. Compounds 1 and 2 were used as templates for the design and synthesis of a series of acyclic immucillin analogues (8-38) in order to identify simplified alternatives to 1 and 2. SerMe-ImmG (8) and DATMe-ImmG (9) displayed the lowest inhibition constants of 2.1 and 3.4 pM, respectively, vs PNP. It was postulated that the flexible natures of 8 and 9 enabled them to adopt conformations resembling those of 1 and 2 within the active site of PNP and that the positioning of two hydroxyl groups was critical for picomolar activity. SerMe-ImmH (10, K(d) = 5.2 pM) was shown to be orally available in mice with a long biological residence time on blood PNP.