Witold Gładkowski

Simultaneous production of citric acid and erythritol from crude glycerol by Yarrowia lipolytica Wratislavia K1

This study shows a possible microbial process for utilization of crude glycerol generated by the biodiesel industry for citric acid and erythritol production. Simultaneous production of citric acid and erythritol under nitrogen-limited conditions with glycerol as the carbon source was achieved with an acetate negative mutant of Y. lipolytica Wratislavia K1 in fed-batch cultivations. The effect of the initial glycerol concentration (from 30–180 g dm−3) on the citrate and erythritol production was investigated. As a result of the experiments, maximum citric acid production (110 g dm−3) and a very high amount of erythritol (81 g dm−3) were determined after 168 h of fed-batch cultivation with the initial glycerol concentration of 150 g dm−3 and the total glycerol concentration of 250 g dm−3. In addition, the citric acid to isocitric acid ratio of the products from this strain was 35.5:1.

Enzymatic enrichment of egg-yolk phosphatidylcholine with α-linolenic acid

Abstractα-Linolenic acid (ALA) was incorporated at 28% into the sn-1 position of egg-yolk phospatidylcholine using Novozyme 435 in one-step transesterification process. Using phospholipase A2 in a two-step process gave 25% incorporation of ALA into the sn-2 position.

An LC method for the analysis of phosphatidylcholine hydrolysis products and its application to the monitoring of the acyl migration process

An assay for quantitative analysis of phosphatidylcholine (1,2-dipalmitoyl-sn-glycero-3-phosphocholine) and its hydrolysis products: 1-hydroxy-2-palmitoyl-sn-glycero-3-phosphocholine and 1-palmitoyl-2-hydroxy-sn-glycero-3-phosphocholine, sn-glycero-3-phosphocholine and palmitic acid using high-performance liquid chromatography with charge aerosol detector (CAD) was developed. The separation of the compounds of interest was achieved on a reversed-phase/hydrophilic interaction stationary phase with a mobile phase consisting of acetonitrile:methanol:10mM ammonium acetate solution. The method was applied to control the acyl migration process of LPC regioisomers in the most common solvents used in the synthesis or modification of PC.

A simple method for positional analysis of phosphatidylcholine.

Simple and fast method of positional analysis of fatty acid composition of phosphatidylcholine (PC) from egg-yolk and soy has been elaborated. The key step of the procedure was complete ethanolysis of PC catalyzed by sn-1,3 specific lipase from Mucor miehei (Lipozyme). 2-Acyl-lysophosphatidylcholine (2-acyl LPC), fatty acids ethyl esters (FAEEs) and free fatty acids (FAs) were formed in this process. No acyl migration was observed during the reaction. The products were entirely separated from the products mixture by simple extraction in water:hexane (2:3 v/v) system. The hexane fraction containing free FAs and FAEEs was treated with BF(3)/Et(2)O in ethanol to obtain only FAEEs. The analysis of FAEEs by GC gave the composition of the FAs in the sn-1 position of the PC. 2-Acyl LPC from water fraction after precipitation in cold (-20°C) acetone was converted into FAEEs and analyzed by gas chromatography (GC) to determine FAs composition in the sn-2 position of the PC.

Lactones 35 [1]. Metabolism of iodolactones with cyclohexane ring in Absidia cylindrospora culture.

The metabolism of δ-iodo-γ-lactones containing cyclohexane ring with an increasing number of methyl substituents in Absidia cylindrospora was studied and seven metabolites were isolated as the products of biotransformations of these substrates. They were formed as the result of various dehalogenation pathways and four of them (hydroxylactones and epoxylactone) turned out to be new compounds. The conversion of substrates ranged from 60% to 90% and the highest conversion was observed for the iodolactone with an unsubstituted cyclohexane ring. The products were fully characterized by the spectroscopic methods and for the hydroxylactone with gem-dimethyl group at C-5 and hydroxylactone with trimethylcyclohexane system the crystal structures were obtained. The main products formed in the process of hydrolytic dehalogenation were δ-hydroxy-γ-lactones with the hydroxy group located cis in relation to lactone moiety. In case of lactone with 4,4,6-trimethylcyclohexane system the dehydrohalogenation followed by the epoxidation of double bond was also observed. One of the metabolites 4,5-epoxy-2,2,6-trimethyl-9-oxabicyclo[4.3.0]nonan-8-one was formed in the sequence of three reactions: hydroxylation at C-5, translactonization and intramolecular nucleophilic substitution of the iodine by the hydroxy group. Some of the isolated products of transformation of the iodolactone with trimethylcyclohexane system were obtained as the single enantiomers. The application of fungi studied to the dehalogenation of iodolactones could be a useful method in the production of new metabolites with oxygen-containing functional groups with antifeedant activity.

Lactones 43. New biologically active lactones: β-cyclocitral derivatives.

BACKGROUND In our previous studies bicyclic γ-lactones with cyclohexane ring exhibited high antifeedant activity against storage pests. The activity was correlated with the type and number of substituents in the cyclohexane ring. One of the most potent group of antifeedant agents was δ-iodo-γ-lactones. RESULTS We present the synthesis of new bicyclic γ-lactones with the cyclohexane ring containing different halogen atoms. To determine the impact of halogen type on biological activity the lactone without halogen atom was also synthesized. The lactones were tested for their antifeedant activity toward the granary weevil beetle (Sitophilus granarius L.), the khapra beetle (Trogoderma granarium Everts) and the confused flour beetle (Tribolium confusum Du Val.). The results of the tests proved that the highest activity was observed for chlorolactone (7) towards larvae and adults of Tribolium confusum. Antibacterial activity of new lactones was also evaluated. Lactone without halogen atom (8) was active against Staphylococcus aureus and Listeria monocytogenes. CONCLUSIONS Studies on the biological activity of synthesised lactones revealed high selectivity towards insect pests as well as bacterial strains. Only the halolactones exhibited significant antifeedant activity. In contrast, antibacterial activity was shown only by the lactone (8) without halogen.

Phosphatidylcholine with cis-9,trans-11 and trans-10,cis-12 Conjugated Linoleic Acid Isomers: Synthesis and Cytotoxic Studies

Novel phosphatidylcholines and lysophosphatidylcholines with cis-9,trans-11 and trans-10,cis-12 conjugated linoleic acid (CLA) were synthesized in high yields (75–99 %). The in vitro cytotoxic activities of these compounds against three human cancer cell lines (HL-60, MCF-7, and HT-29) were evaluated. The results revealed that there are differences in the activity between phosphatidylcholine with cis-9,trans-11 and trans-10,cis-12 CLA acyl groups. 1,2-Di(9Z,11E)-octadecadienoyl-sn-glycero-3-phosphocholine was the most potent cytotoxic agent among all tested CLA derivatives and its IC50 (concentration of a compound that inhibits the proliferation of 50 % of the cancer cell population) was 29.4 µM against HL-60. Moreover, phosphatidylcholines with CLA acyls exhibited much lower cytotoxicity against non-cancer cells (Balb/3T3) than free CLA isomers.

Lactones 46. Synthesis, antifeedant and antibacterial activity of γ‐lactones with a p‐methoxyphenyl substituent

BACKGROUND Lactones are well known for their biological activity. Grosheimin and repin are potent deterrents against storage pests. The unsaturated lactones have exhibited a wide spectrum of antibacterial activity. In our study we focused on the synthesis and evaluation of the biological activity of anisaldehyde derivatives containing lactone function. RESULTS Four new lactones were synthesized in one-step reductive dehalogenation or dehydrohalogenation reactions. These compounds, together with halolactones synthesized earlier, were tested for their antifeedant activity towards Sitophilus granaries, Trogoderma granarium and Tribolium confusum. The results of the tests showed that the highest activity, comparable with that of azadirachtin, towards all tested pests (total coefficient of deterrence 143.3-183.9) was observed for lactone with a vinyl substituent. The antibacterial activity of these compounds was also evaluated. The most potent lactone was active towards gram-positive bacteria strains. CONCLUSIONS The results of biological tests showed that halogen atom removal significantly increased the antifeedant properties of γ-lactones with a p-methoxyphenyl substituent. Unsaturated lactones are most promising in the context of their possible industrial application as crop protection agents. Further structural modifications of lactones with aromatic rings are needed to find important structural factors increasing the antibacterial activity.

Enantioselective Microbial Hydroxylation as a Useful Tool in the Production of Jasmonate Derivatives with Aphid Deterrent Activity.

Microbial transformations of two natural compounds dihydrojasmone (1) and cis-jasmone (3) in the growing cultures of selected twenty strains have been investigated. The studies have demonstrated a biocatalytic potential of tested microorganisms for the enantioselective hydroxylation of jasmonates. The substrates underwent an effective regio- and stero-selective hydroxylation at the allylic position in the cyclopentenone ring, and the corresponding optically pure keto-alcohols (2, 4) were obtained. The process of biohydroxylation depended on the composition of reaction medium. In the studied cultures, (+)-(R)-4-hydroxydihydrojasmone (2) and (+) and (−)-4-hydroxyjasmone (4a, 4b) were produced in good yields and high enantiomeric excesses. Moreover, the introduction of the hydroxy group into the molecule of jasmonate ketones 1 and 3 leads to biologically active derivatives 2 and 4 that regulate the behaviour of aphids Myzus persicae by termination of their feeding.

Isoprenoid-phospholipid conjugates as potential therapeutic agents: Synthesis, characterization and antiproliferative studies

The aim of this research was to extend application field of isoprenoid compounds by their introduction into phospholipid structure as the transport vehicle. The series of novel isoprenoid phospholipids were synthesized in high yields (24–97%), their structures were fully characterized and its anticancer activity was investigated in vitro towards several cell lines of different origin. Most of synthesized compounds showed a significantly higher antiproliferative effect on tested cell lines than free terpene acids. The most active phosphatidylcholine analogue, containing 2,3-dihydro-3-vinylfarnesoic acids instead of fatty acids in both sn-1 and sn-2 position, inhibits the proliferation of colon cancer cells at 13.6 μM.