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Current changes in environmental legislation and customer demands set an urge for the development of more sustainable surfactants. Thus, the objective of this work was the development of novel environmentally friendly amino acid surfactants. Combining Diels–Alder cyclization of myrcene with maleic or citraconic anhydride followed by ring opening with amino acids enabled a synthesis route with a principal 100% atom economy. Variation of amino acids resulted in a large structural variety of anionic and amphoteric surfactants. Lysine gave access to either a mono-acylated product bearing a cationic side chain or a bi-acylated gemini surfactant. First, anhydride precursors were synthesized in yields of >90% in a Diels–Alder reaction under microwave radiation and subsequent amino acid coupling in aqueous environment gave fully bio-based surfactants in good yields and purity. Physicochemical characterization showed an enhanced decrease in surface tension upon addition of amino acids to the myrcene–anhydride backbone, resulting in a minimal value of 31 mN·m−1 for gemini–lysine. Foamabilitiy and foam stability were significantly increased at skin-friendly pH 5.5 by incorporation of amino acids. The carboxylic groups of surfactants with arginine were esterified with ethanol to access cationic compounds. Comparative analysis revealed moderate antimicrobial effects against yeast, Gram-positive bacteria, and Gram-negative bacteria.

Hydroperoxy‐9Z,11E‐octadecadienoic acid (13‐HPODE) can be obtained from safflower oil in an enzyme cascade utilizing lipase, lipoxygenase (LOX), and catalase for in situ oxygen generation. The application of immobilized enzymes may open a new path to a cost‐efficient production of 13‐HPODE, which is used for the synthesis of green note aroma hexanal. Ten immobilization supports are compared for immobilization of lipoxygenase‐1 from Glycine max (LOX‐1) and oxirane‐based Immobead 150 P proves to be best with a maximum LOX‐1 activity of 22 470 U g−1. The immobilizate is successfully recycled in eight consecutive batches and maintains full activity over a period of 16 h using a 3D‐printed column reactor. Catalase from Micrococcus lysodeikticus and LOX‐1 are co‐immobilized on Immobead 150 P allowing a constant production of 13‐HPODE for up to six cycles with a maximum product conversion of 45% and a 13‐regioselectivity of 83%. In this two‐enzyme system with H2O2‐dosage, foam generation is significantly reduced. Co‐immobilization of LOX‐1 and lipaseis possible; however, rapid lipase deactivation occurs. Therefore, a two‐reactor approach with oil hydrolysis in the first reactor is proposed. Immobilized lipases from C. rugosa are suitable for safflower oil hydrolysis and maintain full activity over ten hydrolysis cycles. Practical applications: Linoleic acid hydroperoxide (13‐HPODE) is the starting material for the synthesis of the green note aroma compound hexanal. The byproduct of the hydroperoxide splitting is ω‐oxododecenoic acid, which is currently not employed industrially. The bifunctional oxodocecenoic acid is interesting as precursor for the synthesis of polymer building blocks. Simple one‐step derivatization of the oxo‐group can yield suitable C12 monomers such as dicarboxylic acids, ω‐amino acids, or ω‐hydroxy acids. Cost‐efficiency is a key parameter to incorporate these new biobased building blocks for polymer applications. In this approach, immobilized enzymes are used for the synthesis of 13‐HPODE starting from safflower oil with in situ oxygen generation to prevent excessive foam formation. A two‐reactor concept is designed to circumvent hydroperoxide‐induced lipase deactivation. Direct comparison of both batch and continuous process is performed and provides information for the implementation of the enzyme cascade and the design of an optimized reactor system. [ABSTRACT FROM AUTHOR]

Catechins are the major group of bioactive flavanols in green tea and cacao. 17 glucansucrase-active strains were identified from a set of 41 lactic acid bacteria, which were able to glucosylate (+)-catechin in a non-natural acceptor reaction. In total cell free extracts of 12 Leuconostoc and 5 Weissella strains were active on catechin and also 8 cell fractions exhibited catechin glucosylation activity. Six enzymes were selected for further evaluation and enriched up to 37 fold in yields of at least 40%. Glucansucrase of L. citreum DSM 5577 was the most efficient biocatalyst for (+)-catechin transformation with conversions of >40% after 24 h. NMR analysis of the major reaction product confirmed the (+)-catechin-4′-O-α-d-glucoside. Only L. kimchi B-65337 produced a second catechin monoglucoside. Four out of six glucansucrases glucosylated esculetin and all enzymes were active on haematoxylin. Glucansucrases of L. citreum DSM 5577, L. kimchi B-65337 and W. beninensis DSM 22752 were the best suited biocatalysts with conversions of >30% for esculetin and >60% for haematoxylin. W. beninensis DSM 22752 glucansucrase produced 89% haematoxylin glucosides without process optimization. L. kimchi B-65337 and W. beninensis DSM 22752 synthesized >40% diglucosides with the bifunctional haematoxylin. NMR analysis of the purified esculetin products confirmed formation of the 6-O-α-d- and 7-O-α-d-glucosides. Also two haematoxylin monoglucosides were identified as the 9-O-α-d- and 3-O-α-d-glucosides. [ABSTRACT FROM AUTHOR]

The classical chemical synthesis of CLA triglycerides starting from the free fatty acids and glycerol leads to the formation of additional CLA isomers not suited for applications in the field of human nutrition. Greener methods for a more selective production of CLA triglycerides under gentle reaction conditions were evaluated . The enzymatic synthesis works well under vacuum conditions with a clear preference for free fatty acids as substrates in comparison to alkyl esters. The reaction velocity was enhanced significantly by the addition of basic additives into the reaction mixture. A combined one pot synthesis was designed starting from ethyl esters consisting of an enzymatic ester hydrolysis followed by a re-synthesis of the fatty acids with glycerol. Additionally, a new chemical interesterification reaction with triacetin (triacetyl glyceride) as synthon was developed, which avoids the formation of undesired CLA isomers. [ABSTRACT FROM AUTHOR]

Transketolase A was purified to apparent homogeneity from recombinant Escherichia coli K12 cells carrying the homologous cloned tktA gene on a pUC19-derived plasmid. These recombinant cells exhibited a transketolase activity in crude extracts of up to 9.7 U/mg compared to ≤0.1 U/mg in wild-type cells. Transketolase A was purified from crude extracts of a recombinant strain by successive ammonium sulfate precipitations and two anion-exchange chromatography steps (Q-Sepharose FF, Fractogel EMD-DEAE column) and afforded an apparently homogeneous protein band on SDS/PAGE. The enzyme, both in its active and apoform, had a molecular mass of 145000 Da (± 10000 Da), judged by gel-filtration chromatography. Subunits of 73000 Da ( ± 2000 Da) were determined on SDS/PAGE, thus, transketolase A most likely forms a homodimer. N-terminal amino acid sequencing of the protein verified the identity with the cloned gene tktA. The specific activity of the purified enzyme, determined at 30°C with the substrates xylulose 5-phosphate (donor of C2 compound) and ribose 5-phosphate (acceptor) at an optimal pH (50 mM glycylglycine, pH 8.5), was 50.4 U/mg. Km values for the substrates xylulose 5-phosphate and ribose 5-phosphate were 160 μM and 1.4 mM, respectively. Km values for the other physiological substrates of transketolase A were 90 μM for erythrose 4-phosphate (best acceptor substrate), 2.1 mM for D,L-glyceraldehyde 3-phosphate, 1.1 mM for fructose 6-phosphate, and 4 mM for sedoheptulose 7-phosphate. Hydroxypyruvate served as alternative donor (Km = 18 mM). Unphosphorylated acceptor compounds were formaldehyde (Km = 31 mM), glycolaldehyde (14 mM), D,L-glyceraldehyde (10 mM) and D-erythrose (150 mM). The enzyme was competitively inhibited by D-arabinose 5-phosphate (Ki = 6 mM at a concentration of 2.5 mM D-arabinose 5-phosphate) or by the chelating agent EDTA. The inactive apoform of transketolase A was yielded by dialysis against buffer containing 10 mM EDTA, thus removing the cofactors thiamine diphosphate and divalent cations. The reconstitution of the apoenzyme proceded faster in the presence of manganese ions (Kd = 7 μtM at 10 μM thiamine diphosphate) than with other divalent cations. [ABSTRACT FROM AUTHOR]

Transaldolase catalyzes transfer of a dihydroxyacetone moiety from a ketose donor to an aldose acceptor. During catalysis, a Schiff-base intermediate between dihydroxyacetone and the ϵ-amino group of a lysine residue at the active site of the enzyme is formed. This Schiff-base intermediate has been trapped by reduction with potassium borohydride, and the crystal structure of this complex has been determined at 2.2 Å resolution. The overall structures of the complex and the native enzyme are very similar; formation of the intermediate induces no large conformational changes. The dihydroxyacetone moiety is covalently linked to the side chain of Lys 132 at the active site of the enzyme. The Cl hydroxyl group of the dihydroxyacetone moiety forms hydrogen bonds to the side chains of residues Asn 154 and Ser 176. The C3 hydroxyl group interacts with the side chain of Asp 17 and Asn 35. Based on the crystal structure of this complex a reaction mechanism for transaldolase is proposed. [ABSTRACT FROM AUTHOR]

Starmerella bombicola is known to produce sub‐terminally hydroxylated lactonic sophorolipids (SLs), while Candida kuoi synthesizes acidic open chain SLs with terminally hydroxylated fatty acids. Upon feeding glucose and fatty alcohols both strains form long‐chain nonionic SLs. According to structure elucidation the SLs consist of a hydroxylated fatty acid esterified with fatty alcohol and linked via a glycoside bond to the diacetylated sophorose unit. Palmityl, stearyl, and oleyl alcohols lead to products with lipid chain lengths of C32 or C36. Oleyl alcohol is the preferred substrate leading to 45 g L−1 of the double unsaturated C36 SL with S. bombicola and 20 g L−1 with C. kuoi. Scale up from shake flask to 1.5 L fermentations is possible and 65 g L−1 long‐chain SLs are obtained with S. bombicola within 7 days. Mixed feeding of oleic acid and a variety of fatty alcohols leads to new long‐chain SLs. In the presence of oleic acid the yeasts do not oxidize the fatty alcohol and thus the production of biosurfactants with tailored chain length is possible. The long‐chain SLs show good emulsification ability of water/paraffin oil mixtures at low energy input and reduced interfacial tension significantly. Practical Applications: Sophorolipids are produced by fermentation on industrial scale focusing on cleaning and detergent applications. Mainly lactonic or anionic open‐chain forms are used today. The new long‐chain SLs presented in this manuscript are accessible with existing production technology and can be produced with high titers from cost‐efficient renewable raw materials. In contrast to the commercial products the long‐chain SLs are more hydrophobic and exhibit a strong emulsification behavior. Therefore they have the potential to broaden the application range of SLs in future. They may be useful as novel emulsifiers for cosmetic creams and lotions, pharmaceutical ointments and food products or may find application in oil spill remediation. [ABSTRACT FROM AUTHOR]