The potential of dihydroxyacetone phosphate (DHAP)-dependent aldolases to catalyze stereoselective aldol additions is, in many instances, limited by the solubility of the acceptor aldehyde in aqueous/co-solvent mixtures. Herein, we demonstrate the efficiency of emulsion systems as reaction media for the class I fructose-1,6-bisphosphate aldolase (RAMA) and class II recombinant rhamnulose-1-phosphate aldolase from E. coli (RhuA)-catalyzed aldol addition between DHAP and N-benzyloxycarbonyl (N-Cbz) aminoaldehydes. The use of emulsions improved the RAMA-catalyzed aldol conversions by three to tenfold relative to those in conventional DMF/water mixtures. RhuA was more reactive than RAMA towards the N-Cbz aminoaldehydes regardless of the reaction medium. With (S)- or (R)-Cbz-alaninal, RAMA exhibited preference for the R enantiomer, while RhuA had no enantiomeric discrimination. The linear N-Cbz aminopolyols thus obtained were submitted to catalytic intramolecular reductive amination to afford the corresponding iminocyclitols. This reaction was diastereoselective in all cases examined; the face selectivity was controlled by the stereochemistry of the newly formed hydroxyl group originating from the aldehyde. Characterization of the resulting iminocyclitols allowed the assessment of the diastereoselectivity of the enzymatic aldol reactions with respect to the N-protected aminoaldehyde. RAMA formed single diastereoisomers from N-Cbz-glycinal and from both enantiomers of N-Cbz-alaninal, while 14 % of the epimeric product was observed from N-Cbz-3-aminopropanal. Diastereoselectivity from RhuA was lower than that observed from RAMA. Interestingly, a single diastereoisomer was formed from (S)-Cbz-alaninal, whereas only a 34 % diastereomeric excess was observed from its enantiomer (i.e., (R)-Cbz-alaninal).

The potential of L-fuculose-1-phosphate aldolase (FucA) as a catalyst for the asymmetric aldol addition of dihydroxyacetone phosphate (DHAP) to N-protected amino aldehydes has been investigated. First, the reaction was studied in both emulsion systems and conventional dimethylformamide (DMF)/H2O (1:4 v/v) mixtures. At 100 mM DHAP, compared with the reactions in the DMF/H2O (1:4) mixture, the use of emulsion systems led to two- to three-fold improvements in the conversions of the FucA-catalyzed reactions. The N-protected aminopolyols thus obtained were converted to iminocyclitols by reductive amination with Pd/C. This reaction was highly diastereoselective with the exception of the reaction of the aldol adduct formed from (S)-N-Cbz-alaninal, which gave a 55:45 mixture of both epimers. From the stereochemical analysis of the resulting iminocyclitols, it was concluded that the stereoselectivity of the FucA-catalyzed reaction depended upon the structure of the N-Cbz-amino aldehyde acceptor. Whereas the enzymatic aldol reaction with both enantiomers of N-Cbz-alaninal exclusively gave the expected 3R,4R configuration, the stereochemistry at the C-4 position of the major aldol adducts produced in the reactions with N-Cbz-glycinal and N-Cbz-3-aminopropanal was inverted to the 3R,4S configuration. The study of the FucA-catalyzed addition of DHAP to phenylacetaldehyde and benzyloxyacetaldehyde revealed that the 4R product was kinetically favored, but rapidly disappeared in favor of the 4S diastereoisomer. Computational models were generated for the situations before and after C-C bond formation in the active site of FucA. Moreover, the lowest-energy conformations of each pair of the resulting epimeric adducts were determined. The data show that the products with a 3R,4S configuration were thermodynamically more stable and, therefore, the major products formed, in agreement with the experimental results.

Water-in-oil (W/O) highly concentrated emulsions (gel emulsions) of water/C14E4/aliphatic hydrocarbon systems were investigated as reaction media for the aldolic condensation of dihydroxyacetone phosphate (DHAP) with acceptor aldehydes such as phenylacetaldehyde (1) and benzyloxyacetaldehyde (2), catalyzed by D-fructose-1,6-bisphosphate aldolase from rabbit muscle (RAMA). Prior to any enzymatic reaction, both the formation and stability of the W/O gel emulsions in the presence of reactants were assessed. It was found that the aldehydes improved greatly the kinetic stability of W/O gel emulsions at 25 °C by decreasing the hydrophile-lipophile balance temperature (THLB) of the water/C14E4/aliphatic hydrocarbon systems. Interestingly, the stability of RAMA in W/O gel emulsions was improved by 7-and 25-fold compared to that in aqueous medium or conventional dimethylformamide/water 1/4 v/v mixture, respectively. It was found that the equilibrium yields and enzymatic activity depended on both the aldehyde partitioning between the continuous and dispersed phases and the water-oil interfacial tension. The highest enzymatic activities were achieved in W/O gel emulsion systems with the lowest water-oil interfacial tension. The equilibrium yield depended on the water-oil interfacial tension for the hydrophobic phenylacetaldehyde, and on the partition coefficient for the hydrophilic benzyloxyacetaldehyde. Optimum equilibrium product yields (65-70%) were achieved at either the lowest water-oil interfacial tension or partition coefficient values.

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High-internal-phase-ratio-emulsions (HIPREs) or gel emulsions, formulated with a large amount of water (80.0-99.5% w/w), were investigated as reaction media for α-chymotrypsin-catalysed peptide synthesis under kinetic control using Ac--Phe-OEt and H--Leu-NH 2 as model substrates. Both the initial reaction rate and dipeptide yield were examined as a function of the structure of the non-ionic polyoxyethylene alkyl ether type surfactant, alkyl chain length of the oil component, temperature and aqueous buffer content. Dipeptide yields of 70% were achieved in gel emulsions formulated with 90% w/w aqueous buffer. In these systems, the reaction performance was found to be independent of the gel emulsion system (i.e. surfactant and oil) and therefore of the water-oil interfacial tension. Interestingly, α-chymotrypsin showed superactivity at surfactant concentrations ranging between 0.2 and 0.8% w/w, that is, at 99.5 and 98.0% w/w water content, respectively. Furthermore, high dipeptide yields (90-94%) were achieved in the gel emulsions studied at very high substrate concentrations and thus with undissolved reactants. Under these conditions, examples of α-chymotrypsin-catalysed dipeptide synthesis on an analytical and preparative scale were conducted.

The cover picture shows the doubly charged (q À 2) buta-1,3-diynediyl expanded [3]radialene. The question whether aromaticity enhancement a possible driving force for the facile reduction of some expanded [3]radialenes has been explored. The theoretical study on a series of [3]radialenes with endocyclic and/or peripheral carbo-meric expansion leads to a generalization of the simple 4n2 p z H¸ckel rule to such ™electronic roundabouts∫ with three possible ™exits∫. Thus, the doubly charged (q À 2) buta-1,3-diynediyl expanded [3]radialene shown contains 14 p z electrons in the cross-conjugated ring and exhibits according to the calculations both magnetic and geometrical aromaticity. For more details see the article by R. Chauvin et al. on p. 5056 ff. CONCEPTS ContentsFindings from unexpected reactions were often observed from our attempts to prepare new polynuclear procatalysts and to understand the chemistry of their formation. These could then be applied to syntheses of well-defined heterogeneous olefin polymerization catalysts. We have shown that in the reaction of [Mg 4 (thffo) 8 ] with AlMe 3 the magnesium ± methylalumoxane [Al 3 Mg(m 3 -O)(thffo) 3 (Me) 6 ] (shown here) is formed which is a good methylation agent and support for metallocenes.

Biochemical syntheses O 0035Influence of N-Amino Protecting Group on Aldolase-Catalyzed Aldol Additions of Dihydroxyacetone Phosphate to Amino Aldehydes. -This investigation is presented in extension of recently published aldolase-catalyzed aldol additions of Cbz-protected amino aldehydes. -(CALVERAS, J.; BUJONS, J.; PARELLA, T.; CREHUET, R.; ESPELT, L.; JOGLAR, J.; CLAPES*, P.; Tetrahedron 62 (2006) 11, 2648-2656; Inst. Chem. Environ. Res., CSIC, E-08034 Barcelona, Spain; Eng.) -Nuesgen 26-034